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Sample records for activity calcium deposition

  1. Calcium phosphate/porous silicon biocomposites prepared by cyclic deposition methods: spin coating vs electrochemical activation.

    PubMed

    Hernandez-Montelongo, J; Gallach, D; Naveas, N; Torres-Costa, V; Climent-Font, A; García-Ruiz, J P; Manso-Silvan, M

    2014-01-01

    Porous silicon (PSi) provides an excellent platform for bioengineering applications due to its biocompatibility, biodegradability, and bioresorbability. However, to promote its application as bone engineering scaffold, deposition of calcium phosphate (CaP) ceramics in its hydroxyapatite (HAP) phase is in progress. In that sense, this work focuses on the synthesis of CaP/PSi composites by means of two different techniques for CaP deposition on PSi: Cyclic Spin Coating (CSC) and Cyclic Electrochemical Activation (CEA). Both techniques CSC and CEA consisted on alternate Ca and P deposition steps on PSi. Each technique produced specific morphologies and CaP phases using the same independent Ca and P stem-solutions at neutral pH and at room temperature. The brushite (BRU) phase was favored with the CSC technique and the hydroxyapatite (HAP) phase was better synthesized using the CEA technique. Analyses by elastic backscattering spectroscopy (EBS) on CaP/PSi structures synthesized by CEA supported that, by controlling the CEA parameters, an HAP coating with the required Ca/P atomic ratio of 1.67 can be promoted. Biocompatibility was evaluated by bone-derived progenitor cells, which grew onto CaP/PSi prepared by CSC technique with a long-shaped actin cytoskeleton. The density of adhered cells was higher on CaP/PSi prepared by CEA, where cells presented a normal morphological appearance and active mitosis. These results can be used for the design and optimization of CaP/PSi composites with enhanced biocompatibility for bone-tissue engineering.

  2. Acute calcium pyrophosphate deposition arthropathy.

    PubMed

    Rosen, Thomas; Furman, Janet

    2016-06-01

    Acute calcium pyrophosphate deposition (CPPD) arthropathy, also called pseudogout, is common, and becomes more prevalent as patients age. The presenting symptoms are similar to both gout and septic arthritis but may be treated differently. This article describes a typical patient presentation and management from an emergency medicine and orthopedic surgery standpoint.

  3. Curiosity Finds Calcium-Rich Deposits

    NASA Video Gallery

    NASA’s Curiosity rover finds calcium deposits on Mars similar to thoseseen on Earth when water circulates in cracks and rock fractures.Credit: NASA/JPL-Caltech    › Curiosity's mission site

  4. Calcium Apatite Deposition Disease: Diagnosis and Treatment

    PubMed Central

    2016-01-01

    Calcium apatite deposition disease (CADD) is a common entity characterized by deposition of calcium apatite crystals within and around connective tissues, usually in a periarticular location. CADD most frequently involves the rotator cuff. However, it can theoretically occur in almost any location in the musculoskeletal system, and many different locations of CADD have been described. When CADD presents in an unexpected location it can pose a diagnostic challenge, particularly when associated with pain or swelling, and can be confused with other pathologic processes, such as infection or malignancy. However, CADD has typical imaging characteristics that usually allows for a correct diagnosis to be made without additional imaging or laboratory workup, even when presenting in unusual locations. This is a review of the common and uncommon presentations of CADD in the appendicular and axial skeleton as well as an updated review of pathophysiology of CADD and current treatments. PMID:28042481

  5. The Interaction between Enterobacteriaceae and Calcium Oxalate Deposits

    PubMed Central

    Barr-Beare, Evan; Saxena, Vijay; Hilt, Evann E.; Thomas-White, Krystal; Schober, Megan; Li, Birong; Becknell, Brian; Hains, David S.; Wolfe, Alan J.; Schwaderer, Andrew L.

    2015-01-01

    Background The role of calcium oxalate crystals and deposits in UTI pathogenesis has not been established. The objectives of this study were to identify bacteria present in pediatric urolithiasis and, using in vitro and in vivo models, to determine the relevance of calcium oxalate deposits during experimental pyelonephritis. Methods Pediatric kidney stones and urine were collected and both cultured and sequenced for bacteria. Bacterial adhesion to calcium oxalate was compared. Murine kidney calcium oxalate deposits were induced by intraperitoneal glyoxalate injection and kidneys were transurethrally inoculated with uropathogenic Escherichia coli to induce pyelonephritis Results E. coli of the family Enterobacteriaceae was identified in patients by calcium oxalate stone culture. Additionally Enterobacteriaceae DNA was sequenced from multiple calcium oxalate kidney stones. E. coli selectively aggregated on and around calcium oxalate monohydrate crystals. Mice inoculated with glyoxalate and uropathogenic E. coli had higher bacterial burdens, increased kidney calcium oxalate deposits and an increased kidney innate immune response compared to mice with only calcium oxalate deposits or only pyelonephritis. Conclusions In a murine model, the presence of calcium oxalate deposits increases pyelonephritis risk, likely due to preferential aggregation of bacteria on and around calcium oxalate crystals. When both calcium oxalate deposits and uropathogenic bacteria were present, calcium oxalate deposit number increased along with renal gene transcription of inner stone core matrix proteins increased. Therefore renal calcium oxalate deposits may be a modifiable risk factor for infections of the kidney and urinary tract. Furthermore, bacteria may be present in calcium oxalate deposits and potentially contribute to calcium oxalate renal disease. PMID:26448465

  6. Subcellular localization of calcium deposits during zebrafish (Danio rerio) oogenesis.

    PubMed

    Golpour, Amin; Pšenička, Martin; Niksirat, Hamid

    2016-01-01

    Calcium plays prominent roles in regulating a broad range of physiological events in reproduction. The aim of this study was to describe the subcellular distribution of calcium deposits during stages of oogenesis in zebrafish using a combined oxalate-pyroantimonate technique. The oocyte development of zebrafish was categorized into four stages: primary growth, cortical-alveolus, vitellogenic, and maturation, based on morphological criteria. Calcium deposits in the primary growth stage were detected in the cytoplasm, mitochondria, nucleus, and follicular cells. At the cortical-alveolus stage, calcium particles were transported from follicular cells and deposited in the cortical alveoli. In the vitellogenic stage, some cortical alveoli were compacted and transformed from flocculent electron-lucent to electron-dense objects with the progression of the stage. Calcium deposits were transformed from larger to smaller particles, coinciding with compaction of cortical alveoli. In the maturation stage, calcium deposits in all oocyte compartments decreased, with the exception of those in mitochondria. The proportion of area covered by calcium deposits in the mitochondria and cortical alveoli of oocytes at different stages of development was significantly different (p<0.05). The extent of calcium deposits in the cortical alveoli of mature oocytes was substantially lower than in earlier stages. Basic information about calcium distribution during zebrafish oogenesis may contribute to better understanding of its role in oogenesis.

  7. Ultrastructural localisation of calcium deposits in pig ovarian follicles.

    PubMed

    Rozinek, J; Rajmon, R; Petr, J; Rohlík, J; Jeseta, M; Sedmíková, M; Rehák, D; Jílek, F

    2006-01-01

    Calcium intracellular signaling regulates many intracellular events including oocyte maturation. This signaling is strongly dependent on the influx of calcium ions from extracellular spaces and on the state of intracellular calcium stores. In this study, intracellular calcium deposits were detected in follicle-enclosed pig oocytes using the combined oxalate-pyroantimonate method. These deposits were observed in the nucleus, the mitochondria, the cytoplasm, and on the surface of lipid droplets. The amount of calcium deposits was expressed as a percentage of the area of the respective cellular compartment, which is covered with calcium deposits on ultrathin sections. The distribution of calcium deposits in oocytes changed during folliculogenesis. The amount of calcium deposits in nuclei (1.11% of the area of oocyte nuclei) and cytoplasm (1.02%) in oocytes from secondary and early antral follicles (0.90% nuclei; 0.99% cytoplasm) is significantly lower (P < 0.05) than the amount of calcium deposits in these compartments in oocytes from primary follicles (2.51% nuclei; 2.34% cytoplasm) or antral follicles with growing oocyte (2.91% nuclei; 2.21% cytoplasm). The amount of calcium deposits in mitochondria of oocytes from primary follicles (1.27%) or antral follicles with growing oocyte (1.14%) is significantly lower (P < 0.05) than in the nucleus (2.51% in oocytes from primary follicles; 2.91% in growing oocytes from antral follicles) or cytoplasm (2.34% in oocytes from primary follicles; 2.21% in growing oocytes from antral follicles). The amount of calcium deposits in the cytoplasm of fully-grown oocytes (1.46%) dropped to levels significantly lower (P < 0.05) than those observed in the oocyte nucleus (2.29%). On the basis of these data, we can conclude that the population of follicles on pig ovaries differs in the distribution and concentration of calcium deposits in oocytes, and these changes may be involved in the regulation of the meiotic competence of oocytes.

  8. Stretch-activated calcium channels relay fast calcium waves propagated by calcium-induced calcium influx.

    PubMed

    Jaffe, Lionel F

    2007-03-01

    For nearly 30 years, fast calcium waves have been attributed to a regenerative process propagated by CICR (calcium-induced calcium release) from the endoplasmic reticulum. Here, I propose a model containing a new subclass of fast calcium waves which is propagated by CICI (calcium-induced calcium influx) through the plasma membrane. They are called fast CICI waves. These move at the order of 100 to 1000 microm/s (at 20 degrees C), rather than the order of 3 to 30 microm/s found for CICR. Moreover, in this proposed subclass, the calcium influx which drives calcium waves is relayed by stretch-activated calcium channels. This model is based upon reports from approx. 60 various systems. In seven of these reports, calcium waves were imaged, and, in five of these, evidence was presented that these waves were regenerated by CICI. Much of this model involves waves that move along functioning flagella and cilia. In these systems, waves of local calcium influx are thought to cause waves of local contraction by inducing the sliding of dynein or of kinesin past tubulin microtubules. Other cells which are reported to exhibit waves, which move at speeds in the fast CICI range, include ones from a dozen protozoa, three polychaete worms, three molluscs, a bryozoan, two sea urchins, one arthropod, four insects, Amphioxus, frogs, two fish and a vascular plant (Equisetum), together with numerous healthy, as well as cancerous, mammalian cells, including ones from human. In two of these systems, very gentle local mechanical stimulation is reported to initiate waves. In these non-flagellar systems, the calcium influxes are thought to speed the sliding of actinomyosin filaments past each other. Finally, I propose that this mechanochemical model could be tested by seeing if gentle mechanical stimulation induces waves in more of these systems and, more importantly, by imaging the predicted calcium waves in more of them.

  9. Role of magnesium on the biomimetic deposition of calcium phosphate

    NASA Astrophysics Data System (ADS)

    Sarma, Bimal K.; Sarma, Bikash

    2016-10-01

    Biomimetic depositions of calcium phosphate (CaP) are carried out using simulated body fluid (SBF), calcifying solution and newly developed magnesium containing calcifying solution. Calcium phosphate has a rich phase diagram and is well known for its excellent biocompatibility and bioactivity. The most common phase is hydroxyapatite (HAp), an integral component of human bone and tooth, widely used in orthopedic and dental applications. In addition, calcium phosphate nanoparticles show promise for the targeted drug delivery. The doping of calcium phosphate by magnesium, zinc, strontium etc. can change the protein uptake by CaP nanocrystals. This work describes the role of magnesium on the nucleation and growth of CaP on Ti and its oxide substrates. X-ray diffraction studies confirm formation of HAp nanocrystals which closely resemble the structure of bone apatite when grown using SBF and calcifying solution. It has been observed that magnesium plays crucial role in the nucleation and growth of calcium phosphate. A low magnesium level enhances the crystallinity of HAp while higher magnesium content leads to the formation of amorphous calcium phosphate (ACP) phase. Interestingly, the deposition of ACP phase is rapid when magnesium ion concentration in the solution is 40% of calcium plus magnesium ions concentration. Moreover, high magnesium content alters the morphology of CaP films.

  10. Calcium Activation of Mougeotia Potassium Channels 1

    PubMed Central

    Lew, Roger R.; Serlin, Bruce S.; Schauf, Charles L.; Stockton, Marsha E.

    1990-01-01

    Phytochrome mediates chloroplast movement in the alga Mougeotia, possibly via changes in cytosolic calcium. It is known to regulate a calcium-activated potassium channel in the algal plasma membrane. As part of a characterization of the potassium channel, we examined the properties of calcium activation. The calcium ionophore A23187 activates the channel at external [Ca2+] as low as 20 micromolar. However, external [Ca2+] is not required for activation of the channel by photoactivated phytochrome. Furthermore, when an inhibitor of calcium release from internal stores, 8-(diethylamino)-octyl-3,4,5-trimethoxybenzoate, hydrochloride (TMB-8), is present, red light no longer stimulates channel activity. We conclude that phytochrome activates the plasma membrane potassium channel by releasing calcium from intracellular calcium vesicles; the elevated cytosolic calcium then stimulates channel activity by an unknown mechanism. In the presence of TMB-8, red light does induce chloroplast rotation; thus, potassium channel activation may not be coupled to chloroplast rotation. PMID:16667356

  11. The ANKH gene and familial calcium pyrophosphate dihydrate deposition disease.

    PubMed

    Netter, Patrick; Bardin, Thomas; Bianchi, Arnaud; Richette, Pascal; Loeuille, Damien

    2004-09-01

    Familial calcium pyrophosphate dihydrate deposition (CPPD) disease is a chronic condition in which CPPD microcrystals deposit in the joint fluid, cartilage, and periarticular tissues. Two forms of familial CPPD disease have been identified: CCAL1 and CCAL2. The CCAL1 locus is located on the long arm of chromosome 8 and is associated with CPPD and severe osteoarthritis. The CCAL2 locus has been mapped to the short arm of chromosome 5 and identified in families from the Alsace region of France and the United Kingdom. The ANKH protein is involved in pyrophosphate metabolism and, more specifically, in pyrophosphate transport from the intracellular to the extracellular compartment. Numerous ANKH gene mutations cause familial CCAL2; they enhance ANKH protein activity, thereby elevating extracellular pyrophosphate levels and promoting the formation of pyrophosphate crystals, which produce the manifestations of the disease. Recent studies show that growth factors and cytokines can modify the expression of the normal ANKH protein. These results suggest a role for ANKH in sporadic CPPD disease and in CPPD associated with degenerative disease.

  12. Atomic structure of intracellular amorphous calcium phosphate deposits.

    PubMed

    Betts, F; Blumenthal, N C; Posner, A S; Becker, G L; Lehninger, A L

    1975-06-01

    The radial distribution function calculated from x-ray diffraction of mineralized cytoplasmic structures isolated from the hepatopancreas of the blue crab (Callinectes sapidus) is very similar to that previously found for synthetic amorphous calcium phosphate. Both types of mineral apparently have only short-range atomic order, represented as a neutral ion cluster of about 10 A in longest dimension, whose probable composition is expressed by the formula Ca9(PO4)6. The minor differences observed are attributed to the presence in the biological mineral of significant amounts of Mg-2+ and ATP. Synthetic amorphous calcium phosphate in contact with a solution containing an amount of ATP equivalent to that of the biological mineral failed to undergo conversion to the thermodynamically more stable hydroxyapatite. The amorphous calcium phosphate of the cytoplasmic mineral granules is similarly stable, and does not undergo conversion to hydroxyapatite, presumably owing to the presence of ATP and Mg-2+, known in inhibitors of the conversion process. The physiological implications of mineral deposits consisting of stabilized calcium phosphate ion clusters are discussed.

  13. Boron Induces Early Matrix Mineralization via Calcium Deposition and Elevation of Alkaline Phosphatase Activity in Differentiated Rat Bone Marrow Mesenchymal Stem Cells

    PubMed Central

    Movahedi Najafabadi, Bent-al-hoda; Abnosi, Mohammad Hussein

    2016-01-01

    Objective Boron (B) is essential for plant development and might be an essential micronutrient for animals and humans. This study was conducted to characterize the impact of boric acid (BA) on the cellular and molecular nature of differentiated rat bone marrow mesenchymal stem cells (BMSCs). Materials and Methods In this experimental study, BMSCs were extracted and expanded to the 3rdpassage, then cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) complemented with osteogenic media as well as 6 ng/ml and 6 µg/ml of BA. After 5, 10, 15 and 21 days the viability and the level of mineralization was determined using MTT assay and alizarin red respectively. In addition, the morphology, nuclear diameter and cytoplasmic area of the cells were studied with the help of fluorescent dye. The concentration of calcium, activity of alanine transaminase (ALT), aspartate transaminase (AST), lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) as well as sodium and potassium levels were also evaluated using commercial kits and a flame photometer respectively. Results Although 6 µg/ml of BA was found to be toxic, a concentration of 6 ng/ml increased the osteogenic ability of the cell significantly throughout the treatment. In addition it was observed that B treatment caused the early induction of matrix mineralization compared to controls. Conclusion Although more investigation is required, we suggest the prescription of a very low concentration of B in the form of BA or foods containing BA, in groups at high risk of osteoporosis or in the case of bone fracture. PMID:27054120

  14. Initial deposition of calcium phosphate ceramic on polystyrene and polytetrafluoroethylene by rf magnetron sputtering deposition

    NASA Astrophysics Data System (ADS)

    Feddes, B.; Wolke, J. G. C.; Jansen, J. A.; Vredenberg, A. M.

    2003-03-01

    Calcium phosphate (CaP) coatings can be applied to improve the biological performance of polymeric medical implants. A strong interfacial bond between ceramic and polymer is required for clinical applications. Because the chemical structure of an interface plays an important role in the adhesion of a coating, we studied the formation of the interface between CaP and polystyrene (PS) and polytetrafluoroethylene (PTFE). The coating was deposited in a radio frequency (rf) magnetron sputtering deposition system. Prior to the deposition, some samples received an oxygen plasma pretreatment. We found that the two substrates show a strongly different reactivity towards CaP. On PS a phosphorus and oxygen enrichment is present at the interface. This is understood from POx complexes that are able to bind to the PS. The effects of the plasma pretreatment are overruled by the deposition process itself. On PTFE, a calcium enrichment and an absence of phosphorus is found at the interface. The former is the result of CaF2-like material being formed at the interface. The latter may be the result of phosphorus reacting with escaping fluorine to a PF3 molecule, which than escapes from the material as a gas molecule. We found that the final structure of the interface is mostly controlled by the bombardment of energetic particles escaping either from the plasma or from the sputtering target. The work described here can be used to understand and improve the adhesion of CaP coatings deposited on medical substrates.

  15. Modelling base cations in Europe—sources, transport and deposition of calcium

    NASA Astrophysics Data System (ADS)

    Lee, D. S.; Kingdon, R. D.; Pacyna, J. M.; Bouwman, A. F.; Tegen, I.

    The deposition of the base cations calcium, magnesium and potassium from the atmosphere needs to be quantified in the calculation of the total deposited acidity in the critical loads approach. Of these base cations, calcium has been found to be the most important in terms of mass deposited. However, the sources of calcium to the atmosphere are not well understood. Recently, the first spatially disaggregated inventory of industrial calcium emissions for Europe was presented by Lee and Pacyna (1998) who estimated a total European emission of 0.7-0.8 Mt yr -1. However, it is thought that wind blown dust from soils contributes a substantial fraction to the deposition of calcium. In this work, the source strength of calcium from arid regions within the EMEP modelling domain was estimated using the global mineral dust emission data base of Tegen and Fung (1994) and an estimation of the calcium content of soils. This results in a "natural" calcium emission of 6 Mt yr -1. A long-range transport model, TRACK, was used to calculate the wet and dry deposition of calcium arising from these industrial and natural sources to the UK which resulted in a total deposition of 29-30 kt yr -1. Of this annual deposition, 0.6-0.7 kt arises from cement manufacturing, 0.02-0.03 kt from iron and steel manufacturing, 0.8-0.83 kt from a large point source power generation, and 28 kt from power generation from a small boiler plant. The natural emissions of calcium from arid regions result in a deposition of calcium to the UK of 0.5 kt yr -1. The measured wet deposition of calcium to the UK is 89 kt yr -1 and the estimated dry deposition 14 kt yr -1. The short-fall in the modelled deposition of calcium is thus of the order of 70 kt yr -1, which is suggested to arise from wind-blown dust from agricultural land in the UK and mainland Europe. The estimated emissions, and thus modelled deposition are rather uncertain, such that estimating deposition of calcium attributable to agricultural soil

  16. Computational study of a calcium release-activated calcium channel

    NASA Astrophysics Data System (ADS)

    Talukdar, Keka; Shantappa, Anil

    2016-05-01

    The naturally occurring proteins that form hole in membrane are commonly known as ion channels. They play multiple roles in many important biological processes. Deletion or alteration of these channels often leads to serious problems in the physiological processes as it controls the flow of ions through it. The proper maintenance of the flow of ions, in turn, is required for normal health. Here we have investigated the behavior of a calcium release-activated calcium ion channel with pdb entry 4HKR in Drosophila Melanogaster. The equilibrium energy as well as molecular dynamics simulation is performed first. The protein is subjected to molecular dynamics simulation to find their energy minimized value. Simulation of the protein in the environment of water and ions has given us important results too. The solvation energy is also found using Charmm potential.

  17. Effect of substrate nature on the electrochemical deposition of calcium-deficient hydroxyapatites

    NASA Astrophysics Data System (ADS)

    Gualdrón-Reyes, A. F.; Domínguez-Vélez, V.; Morales-Morales, J. A.; Cabanzo, R.; Meléndez, A. M.

    2017-01-01

    Calcium phosphates were obtained by reducing nitrate ions to produce hydroxide ions on TiO2/stainless steel and TiO2/titanium electrodes. TiO2 coatings on metallic substrates were prepared by sol-gel dip-coating method. The morphology of deposits was observed by FESEM. Chemical nature of calcium phosphate deposits was identified by Raman micro-spectroscopy and FESEM/EDS microanalysis. Electrochemical behavior of nitrate and nitrite reduction on stainless steel and titanium electrodes was studied by linear sweep voltammetry. In addition, voltammetric study of the calcium phosphate electrodeposition on both electrodes was performed. From these measurements was selected the potential to form a calcium phosphate. A catalytic current associated to nitrate reduction reaction was obtained for stainless steel electrode, leading to significant deposition of calcium phosphate. Ca/P ratio for both substrates was less than 1.67. The formation of calcium deficient hydroxyapatite was confirmed by Raman spectroscopy.

  18. Calcium alloy as active material in secondary electrochemical cell

    DOEpatents

    Roche, Michael F.; Preto, Sandra K.; Martin, Allan E.

    1976-01-01

    Calcium alloys such as calcium-aluminum and calcium-silicon, are employed as active material within a rechargeable negative electrode of an electrochemical cell. Such cells can use a molten salt electrolyte including calcium ions and a positive electrode having sulfur, sulfides, or oxides as active material. The calcium alloy is selected to prevent formation of molten calcium alloys resulting from reaction with the selected molten electrolytic salt at the cell operating temperatures.

  19. Calcium and bone disease

    PubMed Central

    Blair, Harry C.; Robinson, Lisa J.; Huang, Christopher L.-H.; Sun, Li; Friedman, Peter A.; Schlesinger, Paul H.; Zaidi, Mone

    2013-01-01

    Calcium transport and calcium signaling are of basic importance in bone cells. Bone is the major store of calcium and a key regulatory organ for calcium homeostasis. Bone, in major part, responds to calcium-dependent signals from the parathyroids and via vitamin D metabolites, although bone retains direct response to extracellular calcium if parathyroid regulation is lost. Improved understanding of calcium transporters and calcium-regulated cellular processes has resulted from analysis of genetic defects, including several defects with low or high bone mass. Osteoblasts deposit calcium by mechanisms including phosphate and calcium transport with alkalinization to absorb acid created by mineral deposition; cartilage calcium mineralization occurs by passive diffusion and phosphate production. Calcium mobilization by osteoclasts is mediated by acid secretion. Both bone forming and bone resorbing cells use calcium signals as regulators of differentiation and activity. This has been studied in more detail in osteoclasts, where both osteoclast differentiation and motility are regulated by calcium. PMID:21674636

  20. Analysis of body calcium (regional changes in body calcium by in vivo neutron activation analysis)

    NASA Technical Reports Server (NTRS)

    Suki, W.; Johnson, P. C.; Leblanc, A.; Evans, H. J.

    1981-01-01

    The effect of space flight on urine and fecal calcium loss was documented during the three long-term Skylab flights. Neutron activation analysis was used to determine regional calcium loss. Various designs for regional analysis were investigated.

  1. Characterization of calcium deposition induced by Synechocystis sp. PCC6803 in BG11 culture medium

    NASA Astrophysics Data System (ADS)

    Yan, Huaxiao; Han, Zuozhen; Zhao, Hui; Zhou, Shixue; Chi, Naijie; Han, Mei; Kou, Xiaoyan; Zhang, Yan; Xu, Linlin; Tian, Chenchen; Qin, Song

    2014-05-01

    Calcium carbonate (CaCO3) crystals in their preferred orientation were obtained in BG11 culture media inoculated with Synechocystis sp. PCC6803 (inoculated BG11). In this study, the features of calcium carbonate deposition were investigated. Inoculated BG11 in different calcium ion concentrations was used for the experimental group, while the BG11 culture medium was used for the control group. The surface morphologies of the calcium carbonate deposits in the experimental and control groups were determined by scanning and transmission electron microscopy. The deposits were analyzed by electronic probe micro-analysis, Fourier transform infrared spectrum, X-ray diffraction, thermal gravimetric analysis and differential scanning calorimetry. The results show that the surfaces of the crystals in the experimental group were hexahedral in a scaly pattern. The particle sizes were micrometer-sized and larger than those in the control group. The deposits of the control group contained calcium (Ca), carbon (C), oxygen (O), phosphorus (P), iron (Fe), copper (Cu), zinc (Zn), and other elements. The deposits in the experimental group contained Ca, C, and O only. The deposits of both groups contained calcite. The thermal decomposition temperature of the deposits in the control group was lower than those in the experimental group. It showed that the CaCO3 deposits of the experimental group had higher thermal stability than those of the control group. This may be due to the secondary metabolites produced by the algae cells, which affect the carbonate crystal structure and result in a close-packed structure. The algae cells that remained after thermal weight loss were heavier in higher calcium concentrations in BG11 culture media. There may be more calcium-containing crystals inside and outside of these cells. These results shall be beneficial for understanding the formation mechanism of carbonate minerals.

  2. Calcium uptake and bioelectrical activity of denervated and myotonic muscle

    PubMed Central

    Radu, H.; Gödri, I.; Albu, E.; Radu, A.; Robu, R.

    1970-01-01

    Calcium uptake on muscle microsomal fraction has been investigated in connection with bioelectrical activity in some muscle diseases. The findings showed a significant increase of calcium uptake in denervated muscle, which exhibited spontaneous bioelectrical activity (fibrillations). In myotonias, a low calcium uptake was peculiar to Steinert's disease but not to myotonia congenita. In other muscle diseases, such as progressive muscular dystrophy (Duchenne's type) or Charcot-Marie-Tooth's disease, the ability of muscle microsomal fraction to bind calcium was not changed. Starting with the key role of calcium in excitation-contraction coupling, the implications of calcium uptake disturbances in muscle electrogenesis are discussed. Images PMID:5431720

  3. Imaging of chondrocalcinosis: calcium pyrophosphate dihydrate (CPPD) crystal deposition disease -- imaging of common sites of involvement.

    PubMed

    Magarelli, N; Amelia, R; Melillo, N; Nasuto, M; Cantatore, F; Guglielmi, G

    2012-01-01

    Calcium pyrophosphate dihydrate (CPPD) crystal deposition disease is characterised by the accumulation of pyrophosphate dihydrate crystals in articular and periarticular tissues and it can be classified as sporadic, hereditary or secondary. The diagnosis frequently rests on radiographic findings. Computed tomography scanning can detect well mineralised deposits in joints and also ultrasound may be useful in detecting CPPD crystal deposits. About MRI recent studies have demonstrated the utility of high field in depiction of CPPD crystal deposits. The aim of this review is to focus on the clinical-classificative and radiological aspects of CPPD, particularly the contribution of the different imaging techniques.

  4. Structures of apicomplexan calcium-dependent protein kinases reveal mechanism of activation by calcium

    SciTech Connect

    Wernimont, Amy K; Artz, Jennifer D.; Jr, Patrick Finerty; Lin, Yu-Hui; Amani, Mehrnaz; Allali-Hassani, Abdellah; Senisterra, Guillermo; Vedadi, Masoud; Tempel, Wolfram; Mackenzie, Farrell; Chau, Irene; Lourido, Sebastian; Sibley, L. David; Hui, Raymond

    2010-09-21

    Calcium-dependent protein kinases (CDPKs) have pivotal roles in the calcium-signaling pathway in plants, ciliates and apicomplexan parasites and comprise a calmodulin-dependent kinase (CaMK)-like kinase domain regulated by a calcium-binding domain in the C terminus. To understand this intramolecular mechanism of activation, we solved the structures of the autoinhibited (apo) and activated (calcium-bound) conformations of CDPKs from the apicomplexan parasites Toxoplasma gondii and Cryptosporidium parvum. In the apo form, the C-terminal CDPK activation domain (CAD) resembles a calmodulin protein with an unexpected long helix in the N terminus that inhibits the kinase domain in the same manner as CaMKII. Calcium binding triggers the reorganization of the CAD into a highly intricate fold, leading to its relocation around the base of the kinase domain to a site remote from the substrate binding site. This large conformational change constitutes a distinct mechanism in calcium signal-transduction pathways.

  5. Calcium Modulation of Plant Plasma Membrane-Bound Atpase Activities

    NASA Technical Reports Server (NTRS)

    Caldwell, C.

    1983-01-01

    The kinetic properties of barley enzyme are discussed and compared with those of other plants. Possibilities for calcium transport in the plasma membrane by proton pump and ATPase-dependent calcium pumps are explored. Topics covered include the ph phase of the enzyme; high affinity of barley for calcium; temperature dependence, activation enthalpy, and the types of ATPase catalytic sites. Attention is given to lipids which are both screened and bound by calcium. Studies show that barley has a calmodulin activated ATPase that is found in the presence of magnesium and calcium.

  6. Calcium pyrophosphate dihydrate deposition disease (CPPD)/Pseudogout of the temporomandibular joint - FNA findings and microanalysis.

    PubMed

    Naqvi, Asghar H; Abraham, Jerrold L; Kellman, Robert M; Khurana, Kamal K

    2008-04-21

    We report a case of a Calcium pyrophosphate dihydrate deposition disease (CPPD) presenting as a mass in the parotid and temporomandibular joint (TMJ) that simulated a parotid tumor. A 35 year-old man presented with pain in the left ear area. A CT Scan of the area showed a large, calcified mass surrounding the left condylar head, and extending into the infratemporal fossa. FNA of the mass showed birefringent crystals, most of which were rhomboid with occasional ones being needle shaped, embedded in an amorphous pink substance. Scanning electron microscopy (SEM) with energy dispersive x-ray spectroscopy (EDS) of these crystals showed peaks corresponding to calcium and phosphorus. SEM/EDS is a rapid method of diagnosing calcium pyrophosphate dihydrate deposition disease (CPPD) and an alternative to more commonly used method of special staining of cell block sections coupled with polarizing microscopy.

  7. Calcium pyrophosphate dihydrate crystal deposition of multiple lumbar facet joints: a case report.

    PubMed

    Namazie, Mohamed Ridzwan bin Mohamed; Fosbender, Murray R

    2012-08-01

    Pseudogout of the lumbar facet joints is rare. We report on a 69-year-old woman with 2-level symptomatic synovial cysts of the facet joints caused by calcium pyrophosphate dihydrate crystal deposition. She underwent surgical decompression for sciatica and low back pain. At one-year follow-up, she had recovered completely.

  8. Hereditary deafness with hydrops and anomalous calcium phosphate deposits

    SciTech Connect

    Johnsson, L.G.; Rouse, R.C.; Hawkins, J.E. Jr.; Kingsley, T.C.; Wright, C.G.

    1981-11-01

    The temporal bones from a 58-year-old white woman who had had hereditary congenital deafness were examined with the techniques of microdissection and surface preparations followed by sectioning of the modiolus. There was bilateral, almost total sensorineural degeneration, which also involved the saccule. The degeneration of the distal processes of the cochlear neurons in the osseous spiral lamina was almost complete, whereas numerous ganglion cells and proximal processes remained in the modiolus and the internal auditory canal. Severe cochleo-saccular hydrops was present in the left ear with Reissner's membrane bulging into the horizontal canal. X-ray diffraction and electron probe analysis were used to study the abnormal crystalline deposits in both ears. On the left side the saccular otoconia were composed of calcite, but the utricular macula was covered by a crust of apatite spherulites. More apatite occurred around the maculae and in the scala media. The cupulae were composed of apatite and octacalcium phosphate. On the right side the utricular otoconia were of normal calcite, but there was a deposit of apatite on the macula sacculi. The upper part of the scala media was completely filled by a deposit of apatite and octacalcium phosphate.

  9. Effects of acid deposition on calcium nutrition and health of Southern Appalachian spruce fir forests

    SciTech Connect

    McLaughlin, S.B.; Wullschleger, S.; Stone, A.; Wimmer, R.; Joslin, J.D.

    1995-02-01

    The role of acid deposition in the health of spruce fir forests in the Southern Appalachian Mountains has been investigated by a wide variety of experimental approaches during the past 10 years. These studies have proceeded from initial dendroecological documentation of altered growth patterns of mature trees to increasingly more focused ecophysiological research on the causes and characteristics of changes in system function associated with increased acidic deposition. Field studies across gradients in deposition and soil chemistry have been located on four mountains spanning 85 km of latitude within the Southern Appalachians. The conclusion that calcium nutrition is an important component regulating health of red spruce in the Southern Appalachians and that acid deposition significantly reduces calcium availability in several ways has emerged as a consistent result from multiple lines or research. These have included analysis of trends in wood chemistry, soil solution chemistry, foliar nutrition, gas exchange physiology, root histochemistry, and controlled laboratory and field studies in which acid deposition and/or calcium nutrition has been manipulated and growth and nutritional status of saplings or mature red spruce trees measured. This earlier research has led us to investigate the broader implications and consequences of calcium deficiency for changing resistance of spruce-fir forests to natural stresses. Current research is exploring possible relationships between altered calcium nutrition and shifts in response of Fraser fir to insect attack by the balsam wooly adelgid. In addition, changes in wood ultrastructural properties in relation to altered wood chemistry is being examined to evaluate its possible role in canopy deterioration, under wind and ice stresses typical of high elevation forests.

  10. Mineral deposition in bacteria-filled and bacteria-free calcium bodies in the crustacean Hyloniscus riparius (Isopoda: Oniscidea).

    PubMed

    Vittori, Miloš; Rozman, Alenka; Grdadolnik, Jože; Novak, Urban; Štrus, Jasna

    2013-01-01

    Crustacean calcium bodies are epithelial sacs which contain a mineralized matrix. The objectives of this study were to describe the microscopic anatomy of calcium bodies in the terrestrial isopod Hyloniscus riparius and to establish whether they undergo molt-related structural changes. We performed 3D reconstruction of the calcium bodies from paraffin sections and analyzed their structure with light and electron microscopy. In addition, we analyzed the chemical composition of their mineralized matrices with micro-Raman spectroscopy. Two pairs of these organs are present in H. riparius. One pair is filled with bacteria while the other pair is not. In non-molting animals, the bacteria-filled calcium bodies contain apatite crystals and the bacteria-free calcium bodies enclose CaCO3-containing concretions with little organic matrix. During preparation for molt, an additional matrix layer is deposited in both pairs of calcium bodies. In the bacteria-filled calcium bodies it contains a mixture of calcium carbonate and calcium phosphate, whereas only calcium carbonate is present in bacteria-free calcium bodies. After ecdysis, all mineral components in bacteria-free calcium bodies and the additional matrix layer in bacteria-filled calcium bodies are completely resorbed. During calcium resorption, the apical surface of the calcium body epithelium is deeply folded and electron dense granules are present in spaces between epithelial cells. Our results indicate that the presence of bacteria might be linked to calcium phosphate mineralization. Calcium bodies likely provide a source of calcium and potentially phosphate for the mineralization of the new cuticle after molt. Unlike other terrestrial isopods, H. riparius does not form sternal CaCO3 deposits and the bacteria-free calcium bodies might functionally replace them in this species.

  11. Effect of calcium cation on lipid vesicle deposition on silicon dioxide surface under various thermal conditions.

    PubMed

    Zhu, Tao; Xu, Fan; Yuan, Bing; Ren, Chunlai; Jiang, Zhongying; Ma, Yuqiang

    2012-01-01

    Calcium cation (Ca(2+)) is a key element to the cell membrane functions. Its effects on liquid crystal vesicle deposition have already been learnt. In this study, it is found that Ca(2+) can also influence the gel vesicle deposition by controlling the vesicle rupture and fusion on SiO(2) at temperatures lower than the main transition temperature. Particular analyses were given to the vesicle-SiO(2) and inter-vesicle attractions that are originated from the Ca(2+) bridging effect. It is concluded that the aggregate condition of vesicles should be taken into consideration when dealing with vesicle deposition on a solid substrate.

  12. Calcium-Activated Phosphate Uptake in Contracting Corn Mitochondria 1

    PubMed Central

    Truelove, B.; Hanson, J. B.

    1966-01-01

    The phosphate inhibition of succinate-powered contraction in corn mitochondria can be reversed with calcium. Associated with this reversal is an accumulation of phosphate and calcium. Both ions are essential for accumulation, although strontium will partially substitute for calcium. Arsenate does not substitute for phosphate except in producing the inhibition of contraction. The antibiotics oligomycin and aurovertin do not block the phosphate inhibition of contraction or the calcium-activated phosphate uptake associated with the release of the inhibition. Dinitrophenol uncouples the phosphate uptake but permits full contraction. Calcium promotes inorganic phosphate accumulation in root tissue as well as in mitochondria. The results are discussed from the viewpoint of theories of calcium reaction with high energy intermediates of oxidative phosphorylation. It is concluded that calcium probably reacts with X∼P in corn mitochondria, rather than with X∼I as with animal mitochondria. PMID:16656343

  13. Evolution of the calcium hydroxyapatite crystal structure under plasma deposition and subsequent reducing treatment

    NASA Astrophysics Data System (ADS)

    Shamrai, V. F.; Karpikhin, A. E.; Sirotinkin, V. P.; Kalita, V. I.; Komlev, D. I.

    2014-03-01

    The structure of hydroxyapatite plasma coatings on a titanium substrate has been investigated by the X-ray Rietveld method. The hydroxyapatite crystal structure in plasma-deposited samples is characterized by strong distortions of its main element (tetrahedral PO4 cluster) and coordination calcium polyhedra, as well as calcium deficit in the Ca2 site; however, these features do not change the main motif of the hydroxyapatite structure. The bond distortions in PO4 clusters are estimated by the Bauer method. It is shown that hydrothermal treatment leads to the almost complete recovery of the hydroxyapatite structure.

  14. Calcium

    MedlinePlus

    ... You'll also find calcium in broccoli and dark green, leafy vegetables (especially collard and turnip greens, ... can enjoy good sources of calcium such as dark green, leafy vegetables, broccoli, chickpeas, and calcium-fortified ...

  15. Increased calcium deposits and decreased Ca2+ -ATPase in erythrocytes of ascitic broiler chickens.

    PubMed

    Li, Kai; Zhao, Lihong; Geng, Guangrui; Ma, Liqin; Dong, Shishan; Xu, Tong; Wang, Jianlin; Wang, Huiyu; Tian, Yong; Qiao, Jian

    2011-06-01

    The decrease of erythrocyte deformability may be one of the predisposing factors for pulmonary hypertension and ascites in broiler chickens. In mammals, the cytoplasmic calcium is a major regulator of erythrocyte deformability. In this study, the erythrocyte deformability was measured, and the precise locations of Ca2+ and Ca2+ -ATPase in the erythrocytes were investigated in chickens with ascites syndrome induced by low ambient temperature. The results showed that ascitic broilers had higher filtration index of erythrocyte compared with control groups, indicating a decrease in erythrocyte deformability in ascitic broilers. The more calcium deposits were observed in the erythrocytes of ascitic broilers compared with those of the age-matched control birds. The Ca2+ -ATPase reactive grains were significantly decreased on the erythrocyte membranes of ascitic broilers. Our data suggest that accumulation of intracellular calcium and inhibition of Ca2+ -ATPase might be important factors for the reduced deformability of the erythrocytes of ascitic broilers.

  16. Subcellular localization of calcium deposits in the noble crayfish Astacus astacus spermatophore: Implications for post-mating spermatophore hardening and spermatozoon maturation.

    PubMed

    Niksirat, Hamid; Kouba, Antonín

    2016-04-01

    The freshly ejaculated spermatophore of crayfish undergoes a hardening process during post-mating storage on the body surface of female. The ultrastructural distribution of calcium deposits were studied and compared in freshly ejaculated and post-mating noble crayfish spermatophores, using the oxalate-pyroantimonate technique, to determine possible roles of calcium in post-mating spermatophore hardening and spermatozoon maturation. Small particles of sparsely distributed calcium deposits were visible in the wall of freshly ejaculated spermatophore. Also, large amount of calcium deposits were visible in the membranes of the freshly ejaculated spermatozoon. Five minutes post-ejaculation, granules in the spermatophore wall appeared as porous formations with numerous electron lucent spaces. Calcium deposits were visible within the spaces and scattered in the spermatophore wall matrix, where smaller calcium deposits combined to form globular calcium deposits. Large numbers of the globular calcium deposits were visible in the wall of the post-mating spermatophore. Smaller calcium deposits were detected in the central area of post-mating spermatophore, which contains the sperm mass, and in the extracellular matrix and capsule. While the density of calcium deposits decreased in the post-mating spermatozoon membranes, numerous small calcium deposits appeared in the subacrosomal zone and nucleus. Substantial changes in calcium deposit distribution in the crayfish spermatophore during post-mating storage on the body of female may be involved in the processes of the spermatophore hardening and spermatozoon maturation.

  17. Antimicrobial Activity of Calcium Hydroxide in Endodontics: A Review

    PubMed Central

    Shalavi, S; Yazdizadeh, M

    2012-01-01

    The purpose of endodontic therapy is to preserve the patient's natural teeth without compromising the patient's local or systemic health. Calcium hydroxide has been included in several materials and antimicrobial formulations that are used in several treatment modalities in endodontics, such as inter-appointment intracanal medicaments. The purpose of this article was to review the antimicrobial properties of calcium hydroxide in endodontics. Calcium hydroxide has a high pH (approximately 12.5-12.8) and is classified chemically as a strong base. The lethal effects of calcium hydroxide on bacterial cells are probably due to protein denaturation and damage to DNA and cytoplasmic membranes. Calcium hydroxide has a wide range of antimicrobial activity against common endodontic pathogens but is less effective against Enterococcus faecalis and Candida albicans. Calcium hydroxide is also a valuable anti-endotoxin agent. However, its effect on microbial biofilms is controversial. PMID:23323217

  18. Mechanisms for the removal of calcium phosphate deposits in turbulent flow

    SciTech Connect

    Littlejohn, F.; Grant, C.S.; Saez, A.E.

    2000-04-01

    This work investigates the mechanisms for the removal of calcium phosphate deposits from stainless steel tubing in turbulent flows. Two types of deposits were analyzed: brushite (dicalcium phosphate dihydrate, DCPD) and a mixture of DCPD/hydroxyapatite (HAP). Cleaning studies were carried out at pHs ranging from 2.85 to 10. The data were analyzed by means of a mathematical model that incorporates the effects of interfacial dissolution and mass transfer. The results show that the HAP/DCPD cleaning rate is influenced both by the kinetics of the interfacial dissolution and by mass transfer. Within the same range of experimental conditions, the rate-limiting mechanism for DCPD removal was the abrasion of the solid by shear stresses. In this case, the interfacial dissolution process plays the role of decreasing the structural integrity of the deposit. These findings show that the removal mechanism of the HAP/DCPD mixture differs significantly from the behavior of individual components.

  19. Calcium Activities During Different Ion Exchange Separation Procedures

    NASA Astrophysics Data System (ADS)

    Zhang, Z.; Zhu, H.; Liu, Y.; Liu, F.; Zhang, C.; Sun, W.

    2014-12-01

    Calcium is a major element and participates in many geological processes. Investigations on stable calcium isotopic compositions of natural geological samples provide a great powerful tool to understand all kinds of those geological processes from a view of the field of isotope geochemistry. With the development of modern instruments and chemical separation techniques, calcium isotopic compositions could be determined even more precisely if the column chemistry brings no deviation. Usually, Calcium is separated from matrix elements using cation resin columns and the related chemical separation techniques seem to be robust. However, more detailed work still need to be done on matrix effects and calcium isotopic fractionations on column chemistry or during elution processes. If calcium is run on TIMS instruments, the interference effect could be lower and easier controlled, thus, the requirement to the chemistry is relatively not critic, but calcium fractionation on filaments could be much difficult to monitor. If calcium is run on MC-ICP-MS instruments, the interference effect could be huge and is really difficult to be recognized and subtracted, the requirement to the chemistry is much more critical in order to get a real result of the sample, but the instrument fractionation could be easier to monitor. Here we investigate calcium activities on several kinds of cation resins under different column/acid conditions. We seek to find a good balance between recovery and interference effect on column chemistry and are intend to set up a better chemical separation procedure to satisfy the instrument requirements for calcium. In addition, Calcium isotopic fractionation on column will also be discussed further here based on our previous and ongoing results.

  20. Synthesis, characterization and osteoblastic activity of polycaprolactone nanofibers coated with biomimetic calcium phosphate.

    PubMed

    Mavis, Bora; Demirtaş, Tolga T; Gümüşderelioğlu, Menemşe; Gündüz, Güngör; Colak, Uner

    2009-10-01

    Immersion of electrospun polycaprolactone (PCL) nanofiber mats in calcium phosphate solutions similar to simulated body fluid resulted in deposition of biomimetic calcium phosphate layer on the nanofibers and thus a highly bioactive novel scaffold has been developed for bone tissue engineering. Coatings with adequate integrity, favorable chemistry and morphology were achieved in less than 6h of immersion. In the coating solutions, use of lower concentrations of phosphate sources with respect to the literature values (i.e., 3.62 vs. 10 mM) was substantiated by a thermodynamic modeling approach. Recipe concentration combinations that were away from the calculated dicalcium phosphate phase stability region resulted in micron-sized calcium phosphates with native nanostructures. While the nano/microstructure formed by the deposited calcium phosphate layer is controlled by increasing the solution pH to above 6.5 and increasing the duration of immersion experimentally, the nanostructure imposed by the dimensions of the fibers was controlled by the polymer concentration (12% w/v), applied voltage (25 kV) and capillary tip to collector distance (35 cm). The deposited coating increased quantitatively by extending the soak up to 6h. On the other hand, the porosity values attained in the scaffolds were around 87% and the biomimetic coatings did not alter the nanofiber mat porosities negatively since the deposition continued along the fibers after the first 2h. Upon confirming the non-toxic nature of the electrospun PCL nanofiber mats, the effects of different nano/microstructures formed were evaluated by the osteoblastic activity. The levels of both alkaline phosphatase activity and osteocalcin were found to be higher in the coated PCL nanofibers than in the uncoated PCL nanofibers, indicating that biomimetic calcium phosphate on PCL nanofibers supports osteoblastic differentiation.

  1. Calcium oxalate deposition in renal allografts: morphologic spectrum and clinical implications.

    PubMed

    Truong, Luan D; Yakupoglu, Ulkem; Feig, Daniel; Hicks, John; Cartwight, Joiner; Sheikh-Hamad, David; Suki, Wadi N

    2004-08-01

    Many aspects of calcium oxalate (CaOx) deposition in renal transplant biopsies are not known. Review of all renal transplant biopsies performed in a 7-year period showed that CaOx deposition could be classified into three groups. Group I: Seven biopsies within a month post-transplant displayed rare CaOx foci against a background of acute tubular necrosis or acute cell-mediated rejection. At follow-up, five grafts functioned well and two failed due to chronic allograft nephropathy. CaOx in this context was an incidental finding secondary to a sudden excretion of an end-stage renal disease-induced increased body burden of CaOx. Group II: Two biopsies performed 2 and 10 months post-transplant showed rare CaOx foci against a background of chronic allograft nephropathy, leading to graft loss. CaOx in this context reflected nonspecific parenchymal deposition due to chronic renal failure regardless of causes. Group III: One biopsy with recurrent PH1 characterized by marked CaOx deposition associated with severe tubulointerstitial injury and graft loss 6 months post-transplant. There were two previously reported cases in which CaOx deposition in the renal allografts was due the antihypertensive drug naftidrofuryl oxalate or increased intestinal absorption of CaOx. CaOx deposition in renal allografts can be classified in different categories with distinctive morphologic features and clinical implications.

  2. Calcium Activation Profile In Electrically Stimulated Intact Rat Heart Cells

    NASA Astrophysics Data System (ADS)

    Geerts, Hugo; Nuydens, Rony; Ver Donck, Luc; Nuyens, Roger; De Brabander, Marc; Borgers, Marcel

    1988-06-01

    Recent advances in fluorescent probe technology and image processing equipment have made available the measurement of calcium in living systems on a real-time basis. We present the use of the calcium indicator Fura-2 in intact normally stimulated rat heart cells for the spatial and dynamic measurement of the calcium excitation profile. After electric stimulation (1 Hz), the activation proceeds from the center of the myocyte toward the periphery. Within two frame times (80 ms), the whole cell is activated. The activation is slightly faster in the center of the cell than in the periphery. The mean recovery time is 200-400 ms. There is no difference along the cell's long axis. The effect of a beta-agonist and of a calcium antagonist is described.

  3. Experimental Comparison of Calcium Sulfate (CaSO(4)) Scale Deposition on Coated Carbon Steel and Titanium Surfaces

    NASA Astrophysics Data System (ADS)

    Al-Otaibi, Dhawi AbdulRahman

    Calcium Sulfate (CaSO4) deposit reduces heat exchange in heat transfer equipment which adversely affects the equipment performance and plant production. This experimental study was conducted by using the Rotating Cylinder Electrode (RCE) equipment available in the university's Center for Engineering Research (CER/RI) to study and compare the effect of solution hydrodynamics on Calcium Sulfate (CaSO4) scale deposition on coated carbon steel and titanium surfaces. In addition, the Scanning Electron Microscopic was used to examine the morphology and distribution of Calcium Sulfate (CaSO 4) crystals deposited on titanium metal surfaces. In this study, the rotational speed was varied from 100 to 2000 RPM to study the behavior of Calcium Sulfate (CaSO4) accumulation on both materials. Based on the experimental results, Calcium Sulfate (CaSO4) scale obtained in the present study was almost constant on coated carbon steel in which the rate of scale deposition is equal to the rate of scale removal. However, the deposition of Calcium Sulfate (CaSO4) observed on titanium material was increased as the speed increased.

  4. Advanced deposition model for thermal activated chemical vapor deposition

    NASA Astrophysics Data System (ADS)

    Cai, Dang

    Thermal Activated Chemical Vapor Deposition (TACVD) is defined as the formation of a stable solid product on a heated substrate surface from chemical reactions and/or dissociation of gaseous reactants in an activated environment. It has become an essential process for producing solid film, bulk material, coating, fibers, powders and monolithic components. Global market of CVD products has reached multi billions dollars for each year. In the recent years CVD process has been extensively used to manufacture semiconductors and other electronic components such as polysilicon, AlN and GaN. Extensive research effort has been directed to improve deposition quality and throughput. To obtain fast and high quality deposition, operational conditions such as temperature, pressure, fluid velocity and species concentration and geometry conditions such as source-substrate distance need to be well controlled in a CVD system. This thesis will focus on design of CVD processes through understanding the transport and reaction phenomena in the growth reactor. Since the in situ monitor is almost impossible for CVD reactor, many industrial resources have been expended to determine the optimum design by semi-empirical methods and trial-and-error procedures. This approach has allowed the achievement of improvements in the deposition sequence, but begins to show its limitations, as this method cannot always fulfill the more and more stringent specifications of the industry. To resolve this problem, numerical simulation is widely used in studying the growth techniques. The difficulty of numerical simulation of TACVD crystal growth process lies in the simulation of gas phase and surface reactions, especially the latter one, due to the fact that very limited kinetic information is available in the open literature. In this thesis, an advanced deposition model was developed to study the multi-component fluid flow, homogeneous gas phase reactions inside the reactor chamber, heterogeneous surface

  5. Reporting neural activity with genetically encoded calcium indicators

    PubMed Central

    Hires, S. Andrew; Tian, Lin; Looger, Loren L.

    2009-01-01

    Genetically encoded calcium indicators (GECIs), based on recombinant fluorescent proteins, have been engineered to observe calcium transients in living cells and organisms. Through observation of calcium, these indicators also report neural activity. We review progress in GECI construction and application, particularly toward in vivo monitoring of sparse action potentials (APs). We summarize the extrinsic and intrinsic factors that influence GECI performance. A simple model of GECI response to AP firing demonstrates the relative significance of these factors. We recommend a standardized protocol for evaluating GECIs in a physiologically relevant context. A potential method of simultaneous optical control and recording of neuronal circuits is presented. PMID:18941901

  6. Eruptive syringomas with calcium deposits in a young woman with Down's syndrome.

    PubMed

    Schepis, C; Torre, V; Siragusa, M; Albiero, F; Cicciarello, R; Gagliardi, M E; Cavallari, V

    2001-01-01

    Eruptive syringomas are uncommon in the general population. We describe here an 18-year-old female, affected by Down's syndrome, who presented with an abrupt eruption of small skin-colored or reddish papules on the face, neck and limbs. Light microscopy allowed us to diagnose syringomas, whereas the study of the ultrastructural features revealed calcium deposits in many lumina and also in the mitochondria. This observation confirms the hypothesis that the syringeal structure plays a role in the pathogenesis of calcinosis cutis.

  7. Biological Origin of Micro-laminated Calcium Carbonate Deposits on Antarctic Rock Surfaces

    NASA Astrophysics Data System (ADS)

    Farmer, M.; Stone, J. O.

    2001-12-01

    We have observed and sampled patchy encrustations of calcium carbonate on rock surfaces in East and West Antarctica. Individual disk-like deposits are up to 1 cm across and a few mm thick, but in places coalesce to form more extensive, colloform coatings. We have observed these deposits on substrates of granite, sandstone, and schist. Their distribution appears similar to that of Antarctic lichens and endolithic algae, extending up to ca. 1000m elevation, but has no consistent relationship to snow drifts, solar radiation, or prevailing winds. The morphology and position of the deposits are distinct from sub-glacial carbonate precipitates. In Marie Byrd Land, the encrustations occur on the surfaces exposed by deglaciation within the past 5000 yrs, and the sample from East Antarctica contains live C-14 (M. Mabin, pers. comm.), suggesting a possible biological origin Electron microprobe and SEM examination of cross-sectioned specimens reveals micron-scale layering of predominantly calcium carbonate, but with a number of bright laminae in SEM images, believed to be calcium fluoride. Sections closely resemble desert varnish in micro-morphology, though not in mineralogy. Isotopic analysis of an organic carbon extract (as opposed to C from the CaCO3 itself) gave a delta C-13 PDB value of -23.3 per mil, similar to values expected in carbon of biological origin. However, we have no proof yet that the carbon analyzed was produced by organisms within the encrustation, rather than being entrapped during an inorganic precipitation process. To investigate the possible biological origin of this material, we attempted to sequence the 16S segment of rRNA in the organic extract, but have not yet completed successful PCR replication. We are continuing attempts to isolate and analyze the pertinent genetic material. The micro-morphology, strongly negative delta C-13 and presence of live C-14 suggest a biological process for precipitation of these calcium carbonate deposits. We hope to

  8. Calcium-dependent Dimerization of Human Soluble Calcium Activated Nucleotidase: Characterization of the Dimer Interface

    SciTech Connect

    Yang,M.; Horii, K.; Herr, A.; Kirley, T.

    2006-01-01

    Mammals express a protein homologous to soluble nucleotidases used by blood-sucking insects to inhibit host blood clotting. These vertebrate nucleotidases may play a role in protein glycosylation. The activity of this enzyme family is strictly dependent on calcium, which induces a conformational change in the secreted, soluble human nucleotidase. The crystal structure of this human enzyme was recently solved; however, the mechanism of calcium activation and the basis for the calcium-induced changes remain unclear. In this study, using analytical ultracentrifugation and chemical cross-linking, we show that calcium or strontium induce noncovalent dimerization of the soluble human enzyme. The location and nature of the dimer interface was elucidated using a combination of site-directed mutagenesis and chemical cross-linking, coupled with crystallographic analyses. Replacement of Ile{sup 170}, Ser{sup 172}, and Ser{sup 226} with cysteine residues resulted in calcium-dependent, sulfhydryl-specific intermolecular cross-linking, which was not observed after cysteine introduction at other surface locations. Analysis of a super-active mutant, E130Y, revealed that this mutant dimerized more readily than the wild-type enzyme. The crystal structure of the E130Y mutant revealed that the mutated residue is found in the dimer interface. In addition, expression of the full-length nucleotidase revealed that this membrane-bound form can also dimerize and that these dimers are stabilized by spontaneous oxidative cross-linking of Cys{sup 30}, located between the single transmembrane helix and the start of the soluble sequence. Thus, calcium-mediated dimerization may also represent a mechanism for regulation of the activity of this nucleotidase in the physiological setting of the endoplasmic reticulum or Golgi.

  9. Activation of purified calcium channels by stoichiometric protein phosphorylation

    SciTech Connect

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

    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.

  10. Cell-compatible properties of calcium carbonates and hydroxyapatite deposited on ultrathin poly(vinyl alcohol)-coated polyethylene films.

    PubMed

    Serizawa, Takeshi; Tateishi, Taishi; Akashi, Mitsuru

    2003-01-01

    Poly(vinyl alcohol) (PVA) was coated onto polyethylene (PE) films by a repetitive adsorption and drying process, and then the PVA-coated PE films were alternately immersed into aqueous solutions of Ca2+ and CO3(2-) ions (alternate soaking cycles), to deposit calcium carbonate (CaCO3) onto the films. The PVA coating was essential for the CaCO3 deposition. The amount of CaCO3 deposited increased with an increasing number of cycles. Scanning electron microscopic observations and attenuated total reflection spectra revealed the presence of both calcite and aragonite as the crystal structures of CaCO3 on the film. L929 fibroblast cells adhered and proliferated on these CaCO3-deposited PE films, as well as the hydroxyapatite-coated PE films previously prepared. It was found that the PVA coating and the subsequent deposition of calcium salts on certain films facilitated cell compatibility.

  11. Mobilization of hepatic calcium pools by platelet activating factor

    SciTech Connect

    Lapointe, D.S.; Hanahan, D.J.; Olson, M.S.

    1987-03-24

    In the perfused rat liver, platelet activating factor, 1-O-hexadecyl-2-acetyl-sn-glycero-3-phosphocholine (AGEPC), infusion produces an extensive but transient glycogenolytic response which at low AGEPC concentrations is markedly dependent upon the perfusate calcium levels. The role of calcium in the glycogenolytic response of the liver to AGEPC was investigated by assessing the effect of AGEPC on various calcium pools in the intact liver. Livers from fed rats were equilibrated with /sup 45/Ca/sup 2 +/, and the kinetics of /sup 45/Ca/sup 2 +/ efflux were determined in control, AGEPC-stimulated, and phenylephrine-stimulated livers during steady-state washout of /sup 45/Ca/sup 2 +/. AGEPC treatment had only a slight if any effect on the pattern of steady-state calcium efflux from the liver, as opposed to major perturbations in the pattern of calcium efflux effected by the ..cap alpha..-adrenergic agonist phenylephrine. Infusion of short pulses of AGEPC during the washout of /sup 45/Ca/sup 2 +/ from labeled livers caused a transient release of /sup 45/Ca/sup 2 +/ which was not abolished at low calcium concentrations in the perfusate. Infusion of latex beads, which are removed by the reticuloendothelial cells, caused the release of hepatic /sup 45/Ca/sup 2 +/ in a fashion similar to the case with AGEPC. The findings indicate that AGEPC does not perturb a major pool of calcium within the liver as occurs upon ..cap alpha..-adrenergic stimulation; it is likely that AGEPC mobilizes calcium from a smaller yet very important pool, very possibly from nonparenchymal cells in the liver.

  12. Calcium imaging of infrared-stimulated activity in rodent brain.

    PubMed

    Cayce, Jonathan Matthew; Bouchard, Matthew B; Chernov, Mykyta M; Chen, Brenda R; Grosberg, Lauren E; Jansen, E Duco; Hillman, Elizabeth M C; Mahadevan-Jansen, Anita

    2014-04-01

    Infrared neural stimulation (INS) is a promising neurostimulation technique that can activate neural tissue with high spatial precision and without the need for exogenous agents. However, little is understood about how infrared light interacts with neural tissue on a cellular level, particularly within the living brain. In this study, we use calcium sensitive dye imaging on macroscopic and microscopic scales to explore the spatiotemporal effects of INS on cortical calcium dynamics. The INS-evoked calcium signal that was observed exhibited a fast and slow component suggesting activation of multiple cellular mechanisms. The slow component of the evoked signal exhibited wave-like properties suggesting network activation, and was verified to originate from astrocytes through pharmacology and 2-photon imaging. We also provide evidence that the fast calcium signal may have been evoked through modulation of glutamate transients. This study demonstrates that pulsed infrared light can induce intracellular calcium modulations in both astrocytes and neurons, providing new insights into the mechanisms of action of INS in the brain.

  13. The Mechanical Environment Modulates Intracellular Calcium Oscillation Activities of Myofibroblasts

    PubMed Central

    Godbout, Charles; Follonier Castella, Lysianne; Smith, Eric A.; Talele, Nilesh; Chow, Melissa L.; Garonna, Adriano; Hinz, Boris

    2013-01-01

    Myofibroblast contraction is fundamental in the excessive tissue remodeling that is characteristic of fibrotic tissue contractures. Tissue remodeling during development of fibrosis leads to gradually increasing stiffness of the extracellular matrix. We propose that this increased stiffness positively feeds back on the contractile activities of myofibroblasts. We have previously shown that cycles of contraction directly correlate with periodic intracellular calcium oscillations in cultured myofibroblasts. We analyze cytosolic calcium dynamics using fluorescent calcium indicators to evaluate the possible impact of mechanical stress on myofibroblast contractile activity. To modulate extracellular mechanics, we seeded primary rat subcutaneous myofibroblasts on silicone substrates and into collagen gels of different elastic modulus. We modulated cell stress by cell growth on differently adhesive culture substrates, by restricting cell spreading area on micro-printed adhesive islands, and depolymerizing actin with Cytochalasin D. In general, calcium oscillation frequencies in myofibroblasts increased with increasing mechanical challenge. These results provide new insight on how changing mechanical conditions for myofibroblasts are encoded in calcium oscillations and possibly explain how reparative cells adapt their contractile behavior to the stresses occurring in normal and pathological tissue repair. PMID:23691248

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

    PubMed Central

    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

  15. Calcium

    MedlinePlus

    ... milligrams) of calcium each day. Get it from: Dairy products. Low-fat milk, yogurt, cheese, and cottage ... lactase that helps digest the sugar (lactose) in dairy products, and may have gas, bloating, cramps, or ...

  16. Initial deposition of calcium phosphate ceramic on polyethylene and polydimethylsiloxane by rf magnetron sputtering deposition: the interface chemistry.

    PubMed

    Feddes, B; Wolke, J G C; Vredenberg, A M; Jansen, J A

    2004-02-01

    Calcium phosphate (CaP) coatings are well known for their bioactive nature. CaP coated polymeric materials can be used as implant material. For this, a strong adhesion between the coating and substrate is necessary. Because the chemical structure of the interface plays an important role in the coating adhesion, we studied the interface between CaP and the polymers polyethylene (PE) and polydimethylsiloxane (PDMS/silicone rubber). Both untreated and plasma pretreated polymers were used. On PE, a low Ca/P ratio nearby the interface and a high amount of C-O bonds were found on both untreated and plasma pretreated PE. This is the result of phosphate-like groups that are able to bind to the carbon of the PE. PDMS reacts towards the plasma pretreatment by losing CH(3) side groups. Compared to PE, a low amount of C-O bonds is found nearby the interface. Besides, a low Ca/P ratio is found nearby the interface. This is the result of phosphate groups that connect to Si atoms of the PDMS, thereby replacing the CH(3) side groups. The bombardment by negatively charged oxygen ions that are accelerated from the target during the deposition process makes the chemical interaction between the coating and the substrates possible.

  17. Engineering of pulsed laser deposited calcium phosphate biomaterials in controlled atmospheres

    NASA Astrophysics Data System (ADS)

    Drukteinis, Saulius E.

    Synthetic calcium phosphates (CAP) such as hydroxyapatite (HA) have been used as regenerative bone graft materials and also as thin films to improve the integration of biomedical implant devices within skeletal tissue. Pulsed laser deposition (PLD) can deposit crystalline HA with significant adhesion on titanium biomaterials. However, there are PLD processing constraints due to the complex physical and chemical interactions occurring simultaneously during PLD, which influence ablation plume formation and development. In this investigation PLD CAP films were engineered with a focus on novel decoupling of partial pressure of H2O (g) ( PH2O ) from total background pressure, in combination with substrate heat treatment and laser energy density control. Characterization of these films was performed with X-ray Diffraction, Scanning Electron Microscopy, Energy Dispersive X-ray Spectroscopy, Fourier Transform Infrared Spectroscopy, and Optical Profilometry. In vitro cellular adhesion testing was also performed using osteoblast (MC3T3) cell lines to evaluate adhesion of bone-forming cells on processed PLD CAP samples. Preferred a-axis orientation films were deposited in H2O (g) saturated atmospheres with reduced laser fluence (< 4 J/cm2). Crystalline HA/tetracalcium phosphate (TTCP) films were deposited in H2O ( g)-deficient atmospheres with higher laser fluence (> 3 J/cm 2). Varied PH2O resulted in control of biphasic HA/TTCP composition with increasing TTCP at lower PH2O . These were dense continuous films composed of micron-scale particles. Cellular adhesion assays did not demonstrate a significant difference between osteoblast adhesion density on HA films compared with biphasic HA/TTCP films. Room temperature PLD at varied PH2O combined with furnace heat treatment resulted in controlled variation in surface amplitude parameters including surface roughness (S a), root mean square (Sq), peak to valley height (St), and ten-point height ( Sz). These discontinuous films were

  18. Decrease in T Cell Activation and Calcium Flux during Clinorotation

    NASA Technical Reports Server (NTRS)

    Sams, Clarence; Holtzclaw, J. David

    2006-01-01

    We investigated the effect of altered gravitational environments on T cell activation. We isolated human, naive T cells (CD3+CD14-CD19-CD16-CD56-CD25-CD69-CD45RA-) following IRB approved protocols. These purified T cells were then incubated with 6 mm polystyrene beads coated with OKT3 (Ortho Biotech, Raritan, NJ) and antiCD28 (Becton Dickinson (BD), San Jose, CA) at 37 C for 24 hours. Antibodies were at a 1:1 ratio and the bead-to-cell ratio was 2:1. Four incubation conditions existed: 1) static or "1g"; 2) centrifugation at 10 relative centrifugal force (RCF) or "10g"; 3) clinorotation at 25 RPM (functional weightlessness or "0g"); and 4) clinorotation at 80 RPM ("1g" plus net shear force approx.30 dynes/sq cm). Following incubation, T cells were stained for CD25 expression (BD) and intracellular calcium (ratio of Fluo4 to Fura Red, Molecular Probes, Eugene, OR) and analyzed by flow cytometry (Coulter EPICS XL, Miami, FL). Results: Static or "1g" T cells had the highest level of CD25 expression and intracellular calcium. T cells centrifuged at 10 RCF ("10g") had lower CD25 expression and calcium levels compared to the static control. However, cells centrifuged at 10 RCF had higher CD25 expression and calcium levels than those exposed to 24 RPM clinorotation ("0g"). T cells exposed to 24 RPM clinorotation had lower CD25 expression, but the approximately the same calcium levels than T cells exposed to 80 RPM clinorotation. These data suggest that stress-activated calcium channel exist in T cells and may play a role during T cell activation.

  19. Hand and wrist arthropathies of hemochromatosis and calcium pyrophosphate deposition disease: distinct radiographic features

    SciTech Connect

    Adamson, T.C. III; Resnik, C.S.; Guerra, J. Jr.; Vint, V.C.; Weisman, M.H.; Resnick, D.

    1983-05-01

    Radiographic features of hand and wrist involvement in 26 patients with hemochromatosis and in 26 patients with idiopathic calcium pyrophosphate dihydrate (CPPD) crystal deposition disease were compared. Two radiologists independently examined the radiographs without knowledge of the specific group to which the patient belonged. The results of this study clearly establish that structural joint diseases in the two disorders are not identical. Characteristic findings allow the radiologist to favor one diagnosis over the other. These radiograhic differences indicate that the arthropathy of hemochromatosis is related to factors additional to the presence of CPPD crystals, specifically, the more prevalent narrowing of the metacarpophalangeal joint spaces, including those in the fourth and fifth digits, peculiar hook-like osteophytes on the radial aspect of the metacarpal heads, and less prevalent separation of the scaphoid and the lunate.

  20. Bone growth on and resorption of calcium phosphate coatings obtained by pulsed laser deposition.

    PubMed

    Clèries, L; Fernández-Pradas, J M; Morenza, J L

    2000-01-01

    Three different calcium phosphate coatings of crystalline hydroxyapatite (HA), alpha- and beta-tricalcium phosphate (alpha+beta-TCP), or amorphous calcium phosphate (ACP) obtained by pulsed laser deposition on Ti-6Al-4V were incubated in a potentially osteogenic primary cell culture (rat bone marrow) in order to evaluate the amount and mode of mineralized bone matrix formation after 2 weeks with special emphasis on the type of interfacial structure that was created. Evaluation techniques included fluorescence labeling and scanning electron microscopy. The resistance to cellular resorption by osteoclasts was also studied. Bone matrix delaminated from the ACP coatings, while it remained on the HA and the alpha+beta-TCP coatings even after fracturing. A cementlike line was seen as the immediate contiguous interface with the nondegrading dense HA surface and with the surface of the remaining porous beta-TCP coating. Highly dense and crystalline HA coatings do not dissolve but are capable of establishing a strong bond with the bone matrix grown on top. Chemical and mechanical bonding were considered in this case. Cellular resorption was practically not observed on the HA coatings, but it was observed on the alpha+beta-TCP coatings. Resorption took place as dissolution that was due to the acidic microenvironment.

  1. Melatonin plus porcine bone on discrete calcium deposit implant surface stimulates osteointegration in dental implants.

    PubMed

    Calvo-Guirado, José Luis; Gómez-Moreno, Gerardo; Barone, Antonio; Cutando, Antonio; Alcaraz-Baños, Miguel; Chiva, Fernando; López-Marí, Laura; Guardia, Javier

    2009-09-01

    The aim of this study was to evaluate the effect of the topical application of melatonin mixed with collagenized porcine bone to accelerate the osteointegration on the rough discrete calcium deposit (DCD) surface implants in Beagle dogs 3 months after their insertion. In preparation for subsequent insertion of dental implants, lower premolars and molars were extracted from 12 Beagle dogs. Each mandible received three parallel wall implants with discrete calcium deposit (DCD) surface of 4 mm in diameter and 10 mm in length. The implants were randomly assigned to the distal sites on each side of the mandible in three groups: group I implants alone, group II implants with melatonin and group III implants with melatonin and porcine bone. Prior to implanting, 5 mg lyophylized powdered melatonin was applied to one bone hole at each side of the mandible. None was applied at the control sites. Ten histological sections per implant were obtained for histomorphometric studies. After a 4-wk treatment period, melatonin significantly increased the perimeter of bone that was in direct contact with the treated implants (P < 0.0001), bone density (P < 0.0001), new bone formation (P < 0.0001) in comparison with control implants. Topical application of melatonin on DCD surface may act as a biomimetic agent in the placement of endo-osseous dental implants and enhance the osteointegration. Melatonin combined with porcine bone on DCD implants reveals more bone to implant contact at 12 wk (84.5 +/- 1.5%) compared with melatonin treated (75.1 +/- 1.4%) and nonmelatonin treated surface implants (64 +/- 1.4%).

  2. Phytase activity from Lactobacillus spp. in calcium-fortified soymilk.

    PubMed

    Tang, Anne Lise; Wilcox, Gisela; Walker, Karen Z; Shah, Nagandra P; Ashton, John F; Stojanovska, Lily

    2010-08-01

    The presence of phytate in calcium-fortified soymilk may interfere with mineral absorption. Certain lactic acid bacteria (LAB) produce the enzyme phytase that degrades phytates and therefore may potentially improve mineral bioavailability and absorption. This study investigates the phytase activity and phytate degradation potential of 7 strains of LAB including: Lactobacillus acidophilus ATCC4962, ATCC33200, ATCC4356, ATCC4161, L. casei ASCC290, L. plantarum ASCC276, and L. fermentum VRI-003. Activity of these bacteria was examined both in screening media and in calcium-fortified soymilk supplemented with potassium phytate. Most strains produced phytase under both conditions with L. acidophilus ATCC4161 showing the highest activity. Phytase activity in fortified soymilk fermented with L. acidophilus ATCC4962 and L. acidophilus ATCC4161 increased by 85% and 91%, respectively, between 12 h and 24 h of fermentation. All strains expressed peak phytase activity at approximately pH 5. However, no phytate degradation could be observed.

  3. Calcium pyrophosphate dihydrate deposition disease (CPPD)/Pseudogout of the temporomandibular joint – FNA findings and microanalysis

    PubMed Central

    Naqvi, Asghar H; Abraham, Jerrold L; Kellman, Robert M; Khurana, Kamal K

    2008-01-01

    We report a case of a Calcium pyrophosphate dihydrate deposition disease (CPPD) presenting as a mass in the parotid and temporomandibular joint (TMJ) that simulated a parotid tumor. A 35 year-old man presented with pain in the left ear area. A CT Scan of the area showed a large, calcified mass surrounding the left condylar head, and extending into the infratemporal fossa. FNA of the mass showed birefringent crystals, most of which were rhomboid with occasional ones being needle shaped, embedded in an amorphous pink substance. Scanning electron microscopy (SEM) with energy dispersive x-ray spectroscopy (EDS) of these crystals showed peaks corresponding to calcium and phosphorus. SEM/EDS is a rapid method of diagnosing calcium pyrophosphate dihydrate deposition disease (CPPD) and an alternative to more commonly used method of special staining of cell block sections coupled with polarizing microscopy. PMID:18426573

  4. {sup 41}Ca as a tracer for calcium uptake and deposition in heart tissue during ischemia and reperfusion

    SciTech Connect

    Southon, J.R.; Bishop, M.S.; Kost, G.J.

    1993-09-17

    We have developed techniques and are commencing experiments using enriched {sup 41}Ca as a tracer in isolated rabbit heart preparations. The aims of the study are to measure calcium uptake and deposition in response to cardiac ischemia and reperfusion, and to investigate events and mechanism leading to irreversible myocyte injury.

  5. Osteoporosis Knowledge, Calcium Intake, and Weight-Bearing Physical Activity in Three Age Groups of Women.

    ERIC Educational Resources Information Center

    Terrio, Kate; Auld, Garry W.

    2002-01-01

    Determined the extent and integration of osteoporosis knowledge in three age groups of women, comparing knowledge to calcium intake and weight bearing physical activity (WBPA). Overall calcium intake was relatively high. There were no differences in knowledge, calcium intake, or WBPA by age, nor did knowledge predict calcium intake and WBPA. None…

  6. The effect of magnetic field on electrochemically deposited calcium phosphate/collagen coatings.

    PubMed

    Zhao, Xueni; He, Jianpeng; Zhang, Jing; Wang, Xudong; Wang, Wanying

    2014-01-01

    Nanostructured calcium phosphate/collagen (CaP/COL) coatings were deposited on the carbon/carbon (C/C) composites through electrochemical deposition (ECD) under magnetic field. The effect of magnetic fields with different orientations on the morphology and composition was investigated. Both the morphology and composition of the coatings could be altered by superimposed magnetic field. Under zero magnetic field and magnetic field, three-dimensional network structure consisting of collagen fibers and CaP were formed on the C/C substrate. The applied magnetic field in the electric field helped to form nanostructured and plate-like CaP on collagen fibers. For the ECD under magnetic field, the Ca/P molar ratio of the coatings was lower than the one under B=0. This may be contributed to the decreased electrical resistance or the increased electrical conductivity of electrolyte solutions under magnetic field. The nanosized CaP/COL coatings exhibited the similar morphology to the human bone and could present excellent cell bioactivity and osteoblast functions.

  7. Determination of fracture toughness of calcium phosphate coatings deposited onto Ti6Al4V substrate by using indentation technique

    NASA Astrophysics Data System (ADS)

    Aydin, Ibrahim; Cetinel, Hakan; Pasinli, Ahmet

    2012-09-01

    In this study, fracture toughness values of calcium phosphate (CaP) coatings deposited onto Ti6Al4V substrate were determined by using Vickers indentation method. In this new patent holding method, the activation processes were performed with NaOH and NaOH+H2O2 on the Ti6Al4V material surface. Thicknesses of CaP coatings were measured from cross-sections of the samples by using optical microscopy. Vickers indentation tests were performed by using microhardness tester. Young's modulus values of the coatings were determined by using ultra microhardness tester. As a result, fracture toughness (K1C) values of the CaP coatings produced by using two different activation processes, were calculated by using experimental study results. These were found to be 0.43 MPa m1/2 and 0.39 MPa m1/2, respectively. It was determined that the CaP coating on Ti6Al4V activated by NaOH+H2O2 had higher fracture toughness than the CaP coating on Ti6Al4V activated by NaOH.

  8. Fractional absorption of active absorbable algal calcium (AAACa) and calcium carbonate measured by a dual stable-isotope method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    With the use of stable isotopes, this study aimed to compare the bioavailability of active absorbable algal calcium (AAACa), obtained from oyster shell powder heated to a high temperature, with an additional heated seaweed component (Heated Algal Ingredient, HAI), with that of calcium carbonate. In ...

  9. Active and passive calcium transport systems in plant cells

    SciTech Connect

    Sze, H.

    1990-01-01

    The ability to change cytoplasmic Ca{sup 2+} levels ((Ca{sup 2+})) by cells has made this cation a key regulator of many biological processes. Cytoplasmic (Ca{sup 2+}) is determined by the coordination of passive Ca{sup 2+} fluxes which increase cytosolic (Ca{sup 2+}) and active Ca{sup 2+} transport systems that lower cytosolic (Ca{sup 2+}). The mechanisms by which plant cells achieve this is poorly understood. We have initially used isolated vesicles from the plasma membrane or organellar membranes to study Ca{sup 2+} transport systems in oat roots (a monocot) and carrot suspension cells (a dicot). The objectives of the proposal were to identify and characterize active (energy-dependent) and passive calcium transport systems that work together to regulate calcium levels in the cytoplasm of plant cells. 10 figs., 2 tabs.

  10. Active and passive calcium transport systems in plant cells

    SciTech Connect

    Sze, H.

    1991-01-01

    The ability to change cytoplasmic Ca{sup 2+} levels ((Ca{sup 2+})) by cells has made this cation a key regulator of many biological processes. Cytoplasmic (Ca{sup 2+}) is determined by the coordination of passive Ca{sup 2+} fluxes which increase cytosolic (Ca{sup 2+}) and active Ca{sup 2+} transport systems that lower cytosolic (Ca{sup 2+}). The mechanisms by which plant cells achieve this is poorly understood. We have initially used isolated vesicles from the plasma membrane or organellar membranes to study Ca{sup 2+} transport systems in oat roots (a monocot) and carrot suspension cells (a dicot). The objectives of the proposal were to identify and characterize active (energy-dependent) and passive calcium transport systems that work together to regulate calcium levels in the cytoplasm of plant cells.

  11. Interaction between Calcium Ions and Bacillus thuringiensis Toxin Activity against Sf9 Cells (Spodoptera frugiperda, Lepidoptera)

    PubMed Central

    Monette, R.; Potvin, L.; Baines, D.; Laprade, R.; Schwartz, J. L.

    1997-01-01

    The effects of calcium ions and modulators of calcium movement on Bacillus thuringiensis insecticidal protein toxicity were investigated with Sf9 cells (Spodoptera frugiperda, fall armyworm) by a new B. thuringiensis toxicity assay based on measurement of fluorescence of ethidium homodimer, a high-affinity DNA stain. CryIC toxicity was substantially stimulated by extracellular calcium in a dose-dependent way (in the millimolar range), while toxicity enhancement could not be replicated when calcium was replaced by barium. This incremental toxicity was reduced by cobalt and lanthanum ions, two inorganic-calcium transport inhibitors. Methoxyverapamil, a voltage-dependent calcium channel blocker, and nifedipine, an inhibitor of dihydropyridine-sensitive L-type calcium channels, had no effect on CryIC toxin activity, but BAY K 8644, an L-type calcium channel activator, increased CryIC activity at high concentrations of extracellular calcium. While A23187, a calcium ionophore, and TMB-8, an inhibitor of intracellular-calcium mobilization, did not change CryIC-induced mortality, thapsigargin, an inhibitor of calcium uptake in intracellular stores, and more particularly trifluoperazine, which inhibits calcium-calmodulin-dependent processes, increased CryIC-mediated toxicity. The incremental effect of extracellular calcium on CryIC-induced toxicity was consistent with an increased concentration of intracellular calcium. PMID:16535509

  12. Calcium phosphates deposited on titanium electrode surface--part 1: Effect of the electrode polarity and oxide film on the deposited materials.

    PubMed

    Okawa, Seigo; Watanabe, Kouichi; Kanatani, Mitsugu

    2013-01-01

    We report experimental results about the effect of polarity of electrode and anodized titanium oxide film on the deposited materials by electrolysis of an acidic calcium phosphate solution. Mirror-polished titanium and anodized titanium were used as anode or cathode, and a Pt plate was used as a counter electrode. The load voltage was held constant at 20 VDC. No deposited materials were found on the anode surface. On the other hand, dicalcium phosphate dihydrate (DCPD) was deposited on the cathode surface at the beginning of the electrolysis. After the electrolysis time 600 s, the non-stoichiometric hydroxyapatite (HAp) with several hundred nanometers was formed on the specimen surface. Based on X-ray photoelectron spectroscopy data, the anodized oxide film contained both P(5+) and P(3+) ions. This characteristic of the oxide film and the electrolysis conditions were related to the behavior of the deposition of ultra fine HAp with high crystallinity.

  13. Synaptosomal calcium influx is activated by sodium fluoride

    SciTech Connect

    Jope, R.S.; Lally, K.M.

    1988-03-15

    Neuronal calcium channels can be modulated by changes in membrane potential or by activation of channel-associated receptors. The latter may be modulated by guanine nucleotide binding proteins. NaF, which activates guanine nucleotide binding proteins, caused a large stimulation of /sup 45/Ca/sup 2 +/ uptake by synaptosomes prepared from rat brain. Stimulation of /sup 45/Ca/sup 2 +/ influx by NaF (i) was apparent in media containing either 5 mM-K+ or 50 mM-K+, (ii) was slower than the fast-phase of voltage-dependent /sup 45/Ca/sup 2 +/ influx but continued for a longer period of time than did depolarization-induced /sup 45/Ca/sup 2 +/ influx, and (iii) was not mimicked or modified by a number of drugs, including ouabain, dinitrophenol, sodium azide or sodium vanadate. These results are consistent with the hypothesis that NaF activates a guanine nucleotide binding protein associated with receptor-coupled calcium channels, resulting in stimulation of calcium influx.

  14. Optogenetic Restoration of Disrupted Slow Oscillations Halts Amyloid Deposition and Restores Calcium Homeostasis in an Animal Model of Alzheimer’s Disease

    PubMed Central

    Kastanenka, Ksenia V.; Hou, Steven S.; Shakerdge, Naomi; Logan, Robert; Feng, Danielle; Wegmann, Susanne; Chopra, Vanita; Hawkes, Jonathan M.; Chen, Xiqun; Bacskai, Brian J.

    2017-01-01

    Slow oscillations are important for consolidation of memory during sleep, and Alzheimer’s disease (AD) patients experience memory disturbances. Thus, we examined slow oscillation activity in an animal model of AD. APP mice exhibit aberrant slow oscillation activity. Aberrant inhibitory activity within the cortical circuit was responsible for slow oscillation dysfunction, since topical application of GABA restored slow oscillations in APP mice. In addition, light activation of channelrhodopsin-2 (ChR2) expressed in excitatory cortical neurons restored slow oscillations by synchronizing neuronal activity. Driving slow oscillation activity with ChR2 halted amyloid plaque deposition and prevented calcium overload associated with this pathology. Thus, targeting slow oscillatory activity in AD patients might prevent neurodegenerative phenotypes and slow disease progression. PMID:28114405

  15. Preparation of calcium-doped boron nitride by pulsed laser deposition

    SciTech Connect

    Anzai, Atsushi; Fuchigami, Masayo; Yamanaka, Shoji; Inumaru, Kei

    2012-08-15

    Highlights: ► Ca-doped boron nitride was prepared by pulsed laser deposition. ► The films do not have long range order structure in terms of XRD. ► But the films had short-range order structure of h-BN sheets. ► Ca-free films had the same optical band gap as crystalline bulk h-BN (5.8 eV.) ► Ca-doping brought about decreases of the optical band gap by ca. 0.4 eV. -- Abstract: Calcium-doped BN thin films Ca{sub x}BN{sub y} (x = 0.05–0.1, y = 0.7–0.9) were grown on α-Al{sub 2}O{sub 3}(0 0 1) substrates by pulsed laser deposition (PLD) using h-BN and Ca{sub 3}N{sub 2} disks as the targets under nitrogen radical irradiation. Infrared ATR spectra demonstrated the formation of short range ordered structure of BN hexagonal sheets, while X-ray diffraction gave no peak indicating the absence of long-range order structure in the films. It was notable that Ca-doped film had 5.45–5.55 eV of optical band gap, while the band gap of Ca-free films was 5.80–5.85 eV. This change in the band gap is ascribed to interaction of Ca with the BN sheets; first principle calculations on h-BN structure indicated that variation of inter-plane distance between the BN layers did not affect the band gap. This study highlights that PLD could prepare BN having short-range structure of h-BN sheets and being doped with electropositive cation which varies the optical band gap of the films.

  16. Deposition and leaching of sulfur, nitrogen and calcium in four forested catchments in China: implications for acidification.

    PubMed

    Larssen, Thorjørn; Duan, Lei; Mulder, Jan

    2011-02-15

    Here we present the first detailed study on fluxes of sulfur (S), nitrogen (N), and major cations in Chinese subtropical forest catchments. Data are from four study sites, differing in inputs of atmospheric pollutants and sensitivity to acidification. Results show important differences from most sites in North America and Europe. Dry deposition of S, N, and calcium (Ca) is considerably larger than wet deposition in most cases causing deposition fluxes ranging from moderate to very high, both for acidifying compounds (S deposition 1.5-10.5 kiloequivalents per hectare and year (keq ha(-1) yr(-1)); N deposition 0.4 to 2.5 keq ha(-1) yr(-1)) and for alkaline compounds (Ca deposition 0.8 to 5.7 keq ha(-1) yr(-1)). More than half of the input of acidity is neutralized by alkalinity associated with Ca deposition. Furthermore, the retention of incoming S and N is small in the soil root zone, but considerable in the deeper soils or riparian zone. Drainage water from the root zone of the soils at the two sites with the highest deposition show pronounced acidification. For the two sites with moderate deposition inputs, the root zones are retaining some of the incoming S and buffer some of the incoming acidity. The subsoils and the riparian zonesare strong sinks for N, S, and Ca. This is associated with substantial acid neutralization at all sites. These features are of major importance for the understanding of the long-term effects of acidification in China.

  17. Calcium and cargoes as regulators of myosin 5a activity

    SciTech Connect

    Sellers, James R. Thirumurugan, Kavitha; Sakamoto, Takeshi; Hammer, John A.; Knight, Peter J.

    2008-04-25

    Myosin 5a is a two-headed actin-dependent motor that transports various cargoes in cells. Its enzymology and mechanochemistry have been extensively studied in vitro. It is a processive motor that takes multiple 36 nm steps on actin. The enzymatic activity of myosin 5 is regulated by an intramolecular folding mechanism whereby its lever arms fold back against the coiled-coil tail such that the motor domains directly bind the globular tail domains. We show that the structure seen in individual folded molecules is consistent with electron density map of two-dimensional crystals of the molecule. In this compact state, the actin-activated MgATPase activity of the molecule is markedly inhibited and the molecule cannot move processively on surface bound actin filaments. The actin-activated MgATPase activity of myosin 5a is activated by increasing the calcium concentration or by binding of a cargo-receptor molecule, melanophilin, in vitro. However, calcium binding to the calmodulin light chains results in dissociation of some of the calmodulin which disrupts the ability of myosin 5a to move on actin filaments in vitro. Thus we propose that the physiologically relevant activation pathway in vivo involves binding of cargo-receptor proteins.

  18. Effect of calcium on adsorption capacity of powdered activated carbon.

    PubMed

    Li, Gang; Shang, Junteng; Wang, Ying; Li, Yansheng; Gao, Hong

    2013-12-01

    We investigated the effect of calcium ion on the adsorption of humic acid (HA) (as a target pollutant) by powered activated carbon. The HA adsorption isotherms at different pH and kinetics of two different solutions including HA alone and HA doped Ca(2+), were performed. It was showed that the adsorption capacity of powdered activated carbon (PAC) for HA was markedly enhanced when Ca(2+) was doped into HA. Also, HA and Ca(2+) taken as nitrate were tested on the uptake of each other respectively and it was showed that the adsorbed amounts of both of them were significantly promoted when HA and calcium co-existed. Furthermore, the adsorbed amount of HA slightly decreased with the increasing of Ca(2+) concentration, whereas the amount of calcium increased with the increasing of HA concentration, but all above the amounts without addition. Finally, the change of pH before and after adsorption process is studied. In the two different solutions including HA alone and HA doped Ca(2+), pH had a small rise, but the extent of pH of later solution was bigger.

  19. ALTERATIONS IN CALCIUM ION ACTIVITY BY ELF AND RF ELECTROMAGNETIC FIELDS

    EPA Science Inventory



    Alterations in calcium ion activity by ELF and RF electromagnetic fields

    Introduction

    Calcium ions play many important roles in biological systems. For example, calcium ion activity can be used as an indicator of second-messenger signal-transduction processe...

  20. Enhancement of enrofloxacin serum antibacterial activity by calcium primed broilers.

    PubMed

    Aguilera, R; Gutiérrez, O L; Sumano, L H

    2007-02-01

    The aim of this trial was to assess the effect that calcium gluconate priming of 468 broilers has on the antibacterial activity of a standard dose of enrofloxacin. Hence, a series of oral pharmacokinetic studies were carried out in four groups of broilers medicated individually through an oral cannula as follows: group A, medicated only with enrofloxacin 10mg/kg; group B, receiving immediately one after the other, calcium gluconate (200mg/kg) and enrofloxacin 10mg/kg; group C, dosed first with calcium gluconate (200mg/kg) and 1h later enrofloxacin (10mg/kg); and group D, dosed first with calcium gluconate (200mg/kg) and 2h later enrofloxacin (10mg/kg). Broilers were bled at different times after the dose of enrofloxacin and antibacterial activity, measured as concentration of enrofloxacin, was measured by an agar diffusion assay. Results revealed that group D the greatest values of maximum serum concentration (Cs(max)), area under the concentration vs. time curve (AUC) and area under the moment curve (AUMC). These values were statistically higher than the corresponding ones derived from groups A, B and C (P<0.05). Taking Cs(max) and AUC values of group A as reference baseline, an increase of 24% and 50%, respectively, was obtained in group D. Group B had the lowest Cs(max), AUC, AUMC and elimination half life (T(1/2)beta) and these values were statistically different from groups A, C and D (P<0.05). The T(1/2)beta was statistically longer in groups C and D as compared with A and B, and the former groups were also different between each other (P<0.05). These results show that if calcium gluconate is first dosed to broilers and 2h later enrofloxacin is administered (as in group D), a more pronounced antibacterial activity of enrofloxacin can be obtained. A challenge of this sequential dosing scheme in a field trial may reveal its clinical value.

  1. Ossifying tendinitis of the rotator cuff after arthroscopic excision of calcium deposits: report of two cases and literature review.

    PubMed

    Merolla, Giovanni; Dave, Arpit C; Paladini, Paolo; Campi, Fabrizio; Porcellini, Giuseppe

    2015-03-01

    Ossifying tendinitis (OT) is a type of heterotopic ossification, characterized by deposition of hydroxyapatite crystals in a histologic pattern of mature lamellar bone. It is usually associated with surgical intervention or trauma and is more commonly seen in Achilles or distal biceps tendons, and also in the gluteus maximus tendon. To our knowledge, there is no description of OT as a complication of calcifying tendinitis of the rotator cuff. In this report, we describe two cases in which the patients developed an OT of the supraspinatus after arthroscopic removal of calcium deposits. The related literature is reviewed.

  2. Lab-Scale Study of the Calcium Carbonate Dissolution and Deposition by Marine Cyanobacterium Phormidium subcapitatum

    NASA Technical Reports Server (NTRS)

    Karakis, S. G.; Dragoeva, E. G.; Lavrenyuk, T. I.; Rogochiy, A.; Gerasimenko, L. M.; McKay, D. S.; Brown, I. I.

    2006-01-01

    Suggestions that calcification in marine organisms changes in response to global variations in seawater chemistry continue to be advanced (Wilkinson, 1979; Degens et al. 1985; Kazmierczak et al. 1986; R. Riding 1992). However, the effect of [Na+] on calcification in marine cyanobacteria has not been discussed in detail although [Na+] fluctuations reflect both temperature and sea-level fluctuations. The goal of these lab-scale studies therefore was to study the effect of environmental pH and [Na+] on CaCO3 deposition and dissolution by marine cyanobacterium Phormidium subcapitatum. Marine cyanobacterium P. subcapitatum has been cultivated in ASN-III medium. [Ca2+] fluctuations were monitored with Ca(2+) probe. Na(+) concentrations were determined by the initial solution chemistry. It was found that the balance between CaCO3 dissolution and precipitation induced by P. subcapitatum grown in neutral ASN III medium is very close to zero. No CaCO3 precipitation induced by cyanobacterial growth occurred. Growth of P. subcapitatum in alkaline ASN III medium, however, was accompanied by significant oscillations in free Ca(2+) concentration within a Na(+) concentration range of 50-400 mM. Calcium carbonate precipitation occurred during the log phase of P. subcapitatum growth while carbonate dissolution was typical for the stationary phase of P. subcapitatum growth. The highest CaCO3 deposition was observed in the range of Na(+) concentrations between 200-400 mM. Alkaline pH also induced the clamping of P. subcapitatum filaments, which appeared to have a strong affinity to envelop particles of chemically deposited CaCO3 followed by enlargement of those particles size. EDS analysis revealed the presence of Mg-rich carbonate (or magnesium calcite) in the solution containing 10-100 mM Na(+); calcite in the solution containing 200 mM Na(+); and aragonite in the solution containing with 400 mM Na(+). Typical present-day seawater contains xxmM Na(+). Early (Archean) seawater was

  3. Influence of precursor solution parameters on chemical properties of calcium phosphate coatings prepared using Electrostatic Spray Deposition (ESD).

    PubMed

    Leeuwenburgh, S C G; Wolke, J G C; Schoonman, J; Jansen, J A

    2004-02-01

    A novel coating technique, referred to as Electrostatic Spray Deposition (ESD), was used to deposit calcium phosphate (CaP) coatings with a variety of chemical properties. The relationship between the composition of the precursor solutions and the crystal and molecular structure of the deposited coatings was investigated by means of X-ray diffraction (XRD), Fourier-Transform Infrared Spectroscopy (FTIR) and Energy Dispersive Spectroscopy (EDS). It was shown that the relative Ca/P ratio in the precursor solution, the absolute precursor concentration, the acidity of the precursor solution and the type of Ca-precursor strongly influenced the chemical nature of the deposited CaP coatings. Various crystal phases and phase mixtures were obtained, such as carbonate apatite, beta-TCP, Mg-substituted whitlockite, monetite, beta/gamma-pyrophosphate, and calcite. It was shown that carbonate plays an essential role in the chemical mechanism of coating formation. Carbonate is formed due to a decomposition reaction of organic solvents. Depending on deposition conditions, carbonate anions (a) react with acidic phosphate groups, (b) are incorporated into apatitic calcium phosphate phases, and (c) react with excessive Ca(2+) cations in case of phosphate-deficient precursor solutions.

  4. Response of sediment calcium and magnesium species to the regional acid deposition in eutrophic Taihu Lake, China.

    PubMed

    Tao, Yu; Dan, Dai; Chengda, He; Qiujin, Xu; Fengchang, Wu

    2016-11-01

    Acid deposition causes carbonate dissolution in watersheds and leads to profound impacts on water chemistry of lakes. Here, we presented a detailed study on the seasonal, spatial, and vertical variations of calcium and magnesium species in the overlying water, interstitial water, and sediment profiles in eutrophic Taihu Lake under the circumstance of regional acid deposition. The result showed that both the acid deposition and biomineralization in Taihu Lake had effects on Ca and Mg species. In the lake water, calcium carbonate was saturated or over-saturated based on long-term statistical calculation of the saturation index (SI). On the sediment profiles, significant difference in Ca and Mg species existed between the surface sediment (0-10 cm) and deeper sediments (>10 cm). The interstitial water Ca(2+) and Mg(2+), ion-exchangeable Ca and Mg in the surface sediment were higher than those in the deeper sediment. In the spring, when the acid deposition is more intensive, the acid-extracted Ca and Mg in the surface sediment were lower than that in the deeper sediment in the northwest lake, due to carbonate dissolution caused by the regional acid deposition. Spatially, the higher concentration of acid-extracted Ca and Mg in the northwest surface sediment than that in the east lake was observed, indicating the pronounced carbonate biomineralization by algae bloom in the northwest lake. Statistical analysis showed that acid deposition exerted a stronger impact on the variation of acid-extracted Ca and Mg in the surface sediment than the biomineralization in Taihu Lake. For the total Ca and Mg concentration in the spring, however, no significant change between the surface and deeper sediment in the northwest lake was observed, indicating that the carbonate precipitation via biomineralization and the carbonate dissolution due to acidic deposition were in a dynamic balance. These features are of major importance for the understanding of combined effects of acid

  5. Responses in calcium and phosphorus metabolism and hepatic lipid deposition among estrogenized chicks fed various dietary ingredients.

    PubMed

    Bolden, S L; Jensen, L S; Takahashi, K

    1984-03-01

    The purpose of this study was to determine whether diet composition would influence calcium and phosphorus metabolism in chicks administered estrogen. At 1 day of age, broiler chicks were fed either a corn-soybean meal diet (CS), or an isoenergetic and isonitrogenous diet containing 5% fish meal, 5% alfalfa meal and 10% torula yeast (FAY). At 21 days equivalent numbers were implanted with one of two lengths of Silastic tubing containing estradiol dipropionate, while the remaining birds served as nonimplanted controls. Significant increases were observed in liver weight, liver lipid, plasma total calcium and inorganic phosphate in chicks that were implanted, while concomitant declines were seen in body weight. Implanted chicks fed the CS diet had significantly higher liver weight, liver lipid, plasma phosphorus and plasma calcium and lower tibial bone ash than those fed the FAY diet. Furthermore, liver lipid values were very highly correlated with plasma phosphorus and calcium. In an identical study with slower growing White Leghorn chicks, the same trends were observed but were less well defined. These data show that the inclusion of certain ingredients into corn-soybean diets balanced for the major nutrients affects the response of chicks to estrogenization. Liver lipid deposition, calcium and phosphorus metabolism are all subject to diet and estrogen interactions.

  6. Synthesis of Calcium(II) Amidinate Precursors for Atomic Layer Deposition through a Redox Reaction between Calcium and Amidines.

    PubMed

    Kim, Sang Bok; Yang, Chuanxi; Powers, Tamara; Davis, Luke M; Lou, Xiabing; Gordon, Roy G

    2016-08-22

    We have prepared two new Ca(II) amidinates, which comprise a new class of ALD precursors. The syntheses proceed by a direct reaction between Ca metal and the amidine ligands in the presence of ammonia. Bis(N,N'-diisopropylformamidinato)calcium(II) (1) and bis(N,N'-diisopropylacetamidinato)calcium(II) (2) adopt dimeric structures in solution and in the solid state. X-ray crystallography revealed asymmetry in one of the bridging ligands to afford the structure [(η(2) -L)Ca(μ-η(2) :η(2) -L)(μ-η(2) :η(1) -L)Ca(η(2) -L)]. These amidinate complexes showed unprecedentedly high volatility as compared to the widely employed and commercially available Ca(II) precursor, [Ca3 (tmhd)6 ]. In CaS ALD with 1 and H2 S, the ALD window was approximately two times wider and lower in temperature by about 150 °C than previously reported with [Ca3 (tmhd)6 ] and H2 S. Complexes 1 and 2, with their excellent volatility and thermal stability (up to at least 350 °C), are the first homoleptic Ca(II) amidinates suitable for use as ALD precursors.

  7. Synthesis of Calcium(II) Amidinate Precursors for Atomic Layer Deposition through a Redox Reaction between Calcium and Amidines

    PubMed Central

    Kim, Sang Bok; Yang, Chuanxi; Powers, Tamara; Davis, Luke M.; Lou, Xiabing

    2016-01-01

    Abstract We have prepared two new CaII amidinates, which comprise a new class of ALD precursors. The syntheses proceed by a direct reaction between Ca metal and the amidine ligands in the presence of ammonia. Bis(N,N′‐diisopropylformamidinato)calcium(II) (1) and bis(N,N′‐diisopropylacetamidinato)calcium(II) (2) adopt dimeric structures in solution and in the solid state. X‐ray crystallography revealed asymmetry in one of the bridging ligands to afford the structure [(η2‐L)Ca(μ‐η2:η2‐L)(μ‐η2:η1‐L)Ca(η2‐L)]. These amidinate complexes showed unprecedentedly high volatility as compared to the widely employed and commercially available CaII precursor, [Ca3(tmhd)6]. In CaS ALD with 1 and H2S, the ALD window was approximately two times wider and lower in temperature by about 150 °C than previously reported with [Ca3(tmhd)6] and H2S. Complexes 1 and 2, with their excellent volatility and thermal stability (up to at least 350 °C), are the first homoleptic CaII amidinates suitable for use as ALD precursors. PMID:27351794

  8. Toxic industrial deposit remediation by ant activity

    NASA Astrophysics Data System (ADS)

    Jilkova, Veronika; Frouz, Jan

    2016-04-01

    Toxic industrial deposits are often contaminated by heavy metals and the substrates have low pH values. In such systems, soil development is thus slowed down by high toxicity and acidic conditions which are unfavourable to soil fauna. Ants (Hymenoptera, Formicidae) are considered tolerant to heavy metal pollution and are known to increase organic matter content and microbial activity in their nests. Here, we focused on soil remediation caused by three ant species (Formica sanguinea, Lasius niger, and Tetramorium sp.) in an ore-washery sedimentation basin near Chvaletice (Czech Republic). Soil samples were taken from the centre of ant nests and from the nest surroundings (>3 m from nests). Samples were then analyzed for microbial activity and biomass and contents of organic matter and nutrients. As a result, ant species that most influenced soil properties was F. sanguinea as there were higher microbial activity and total nitrogen and ammonia contents in ant nests than in the surrounding soil. We expected such a result because F. sanguinea builds conspicuous large nests and is a carnivorous species that brings substantial amounts of nitrogen in insect prey to their nests. Effects of the other two ant species might be lower because of smaller nests and different feeding habits as they rely mainly on honeydew from aphids or on plant seeds that do not contain much nutrients.

  9. Systematic review and quality analysis of emerging diagnostic measures for calcium pyrophosphate crystal deposition disease

    PubMed Central

    Wu, Y; Chen, K; Terkeltaub, R

    2016-01-01

    Objectives Calcium pyrophosphate crystal deposition disease (CPPD) is common, yet prevalence and overall clinical impact remain unclear. Sensitivity and specificity of CPPD reference standards (conventional crystal analysis (CCA) and radiography (CR)) were meta-analysed by EULAR (published 2011). Since then, new diagnostic modalities are emerging. Hence, we updated 2009–2016 literature findings by systematic review and evidence grading, and assessed unmet needs. Methods We performed systematic search of full papers (PubMed, Scopus/EMBASE, Cochrane 2009–2016 databases). Search terms included CPPD, chondrocalcinosis, pseudogout, ultrasound, MRI, dual energy CT (DECT). Paper selection, data abstraction, EULAR evidence level, and Quality Assessment of Diagnostic Accuracy Studies (QUADAS)-2 bias and applicability grading were performed independently by 3 authors. Results We included 26 of 111 eligible papers, which showed emergence in CPPD diagnosis of ultrasound (U/S), and to lesser degree, DECT and Raman spectroscopy. U/S detected CPPD crystals in peripheral joints with sensitivity >80%, superior to CR. However, most study designs, though analytical, yielded low EULAR evidence level. DECT was marginally explored for CPPD, compared with 35 published DECT studies in gout. QUADAS-2 grading indicated strong applicability of U/S, DECT and Raman spectroscopy, but high study bias risk (in ∼30% of papers) due to non-controlled designs, and non-randomised subject selection. Conclusions Though CCA and CR remain reference standards for CPPD diagnosis, U/S, DECT and Raman spectroscopy are emerging U/S sensitivity appears to be superior to CR. We identified major unmet needs, including for randomised, blinded, controlled studies of CPPD diagnostic performance and rigorous analyses of 4 T MRI and other emerging modalities. PMID:27933211

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

    NASA Technical Reports Server (NTRS)

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

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

  11. Calcium dependence and distribution of calcium-activated chloride channels in Xenopus oocytes.

    PubMed Central

    Gomez-Hernandez, J M; Stühmer, W; Parekh, A B

    1997-01-01

    1. The Ca(2+)-dependent Cl- current (ICl,Ca), expressed in the plasma membrane of Xenopus oocytes, was examined in excised inside-out macropatches using a rapid perfusion system. 2. Application of Ca(2+)-containing Ringer solution resulted in the activation of a current whose reversal potential shifted to the right by 51 +/- 5.2 mV when Cl- in the pipette solution was lowered from 119.3 to 10 mM. No currents were generated when Ca2+ was omitted from the solution. The current is therefore a Ca(2+)-activated Cl- one. 3. Following exposure to Ca2+, the half-time for activation of ICl,Ca was not voltage dependent, whereas deactivation was strongly so. 4. ICl,Ca was stable in the continuous presence of Ca2+ and showed no sign of inactivation or adaptation. 5. Comparison of the size of the currents (normalized to pipette resistance) from the animal and vegetal poles revealed that ICl,Ca had a highly polarized distribution. The current density was almost 10 times higher in the animal pole. 6. The results suggest that Cl- channels provide a continuous and reliable indication of submembranous Ca2+, at least in an excised patch, and the clustering of the Cl- channels renders it necessary to exert caution in interpreting results involving the kinetics of Ca2+ signalling, when ICl,Ca is used as the sole monitor of calcium. PMID:9279809

  12. Effect of calcium on the hemolytic activity of Stichodactyla helianthus toxin sticholysin II on human erythrocytes.

    PubMed

    Celedón, Gloria; González, Gustavo; Lissi, Eduardo; Cerda, Tania; Martinez, Diana; Soto, Carmen; Pupo, Mario; Pazos, Fabiola; Lanio, Maria E; Alvarez, Carlos

    2009-11-01

    Sticholysin II (St II) is a toxin from the sea anemona Stichodactyla helianthus that produces erythrocytes lysis at low concentration and its activity depends on the presence of calcium. Calcium may act modifying toxin interaction with erythrocyte membranes or activating cellular processes which may result in a modified St II lytic action. In this study we are reporting that, in the presence of external K(+), extracellular calcium decreased St II activity on erythrocytes. On the other hand an increase of intracellular calcium promotes Sty II lytic activity. The effect of intracellular calcium was specifically studied in relation to membrane lipid translocation elicited by scramblases and how this action influence St II lytic activity on erythrocytes. We used 0.5 mmol/L calcium and 10 mmol/L A23187, as calcium ionophore, for scramblases activation and found increased St II activity associated to increase of intracellular calcium. N-ethyl maleimide (activator) and 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (inhibitor) were used as scramblases modulators in the assays which produced an increase and a decrease of the calcium effect, respectively. Results reported suggest an improved St II membrane pore-forming capacity promoted by intracellular calcium associated to membrane phospholipids translocation.

  13. Microdamage induced calcium efflux from bone matrix activates intracellular calcium signaling in osteoblasts via L-type and T-type voltage-gated calcium channels.

    PubMed

    Jung, Hyungjin; Best, Makenzie; Akkus, Ozan

    2015-07-01

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

  14. Low temperature solution deposition of calcium phosphate coatings for orthopedic implants

    SciTech Connect

    Campbell, A.A.; Graff, G.L.

    1994-04-01

    Calcium phosphate coatings were grown from aqueous solution onto a derivatized self-assmebled monolayer (SAM) which was covalently bound to a titanium metal substrate. The SAM molecules provided an idea connection between the metal surface and the calcium phosphate coating. The trichlorosilane terminus of the SAM molecule insured covalent attachment to the surface, while the functionalized ``tail`` induced heterogeneous nucleation of the calcium phosphate coating from supersaturated solutions. This low temperature process allowed for uniform coatings to be produced onto complex-shaped and/or microporous surfaces and provided better control of phase purity.

  15. Migration of calcium deposits into subacromial-subdeltoid bursa and into humeral head as a rare complication of calcifying tendinitis: sonography and imaging.

    PubMed

    Della Valle, Valeria; Bassi, Emilio Maria; Calliada, Fabrizio

    2015-09-01

    Calcifying tendinitis of the shoulder is a common condition characterized by the deposition of calcium, predominantly hydroxyapatite crystals, in the rotator cuff. A rare complication of this condition is the migration of calcium deposits from tendons, usually the supraspinatus, into the subacromial-subdeltoid bursa or into the humeral greater tuberosity. These complications are responsible for intense acute shoulder pain and functional disability. Patient anamnesis and clinical symptoms must be considered to make the diagnosis, but imaging, particularly sonography, is often necessary, showing a typical presentation related to the locations of calcium deposits. We present sonographic and other imaging features of subacromial-subdeltoid bursitis and humeral osteitis related to the migration of calcium.

  16. Bioactive calcium phosphate coating formed on micro-arc oxidized magnesium by chemical deposition

    NASA Astrophysics Data System (ADS)

    Liu, G. Y.; Hu, J.; Ding, Z. K.; Wang, C.

    2011-01-01

    In order to improve the bioactivity of the micro-arc oxidized magnesium, a calcium phosphate coating was formed on the surface of micro-arc oxidized magnesium using a chemical method. The microstructures of the substrate and the calcium phosphate coating before and after the simulated body fluids (SBF) incubation were characterized by X-ray diffraction, Fourier-transformed infrared spectroscopy and scanning electron microscopy. The results showed that the calcified coating was composed of calcium deficient hydroxyapatite (HA) and dicalcium phosphate dihydrate (DCPD). After SBF incubation, some new apatite formation on the calcified coating surface from SBF could be found. The corrosion behaviours of the samples in SBF were also investigated by potentiodynamic polarization curves and immersion tests. The results showed that calcium phosphate coating increased the corrosion potential, and decreased the hydrogen gas release.

  17. Anti-melanin deposition activity of ceramicines from Chisocheton ceramicus.

    PubMed

    Iijima, Chie; Wong, Chin Piow; Nugroho, Alfarius Eko; Sotozono, Yayoi; Someya, Saki; Hirasawa, Yusuke; Kaneda, Toshio; Hadi, A Hamid A; Morita, Hiroshi

    2016-10-01

    The ceramicines, a series of limonoids from Chisocheton ceramicus (Meliaceae), were evaluated for anti-melanin deposition activity on α-melanocyte stimulating hormone (α-MSH) and 3-isobutyl-1-methylxanthine (IBMX)-treated B16-F10 melanoma cell, and several ceramicines were found to be active. The structure-activity relationship of ceramicines as anti-melanin deposition inhibitors was deduced. Furthermore, the mechanism of anti-melanin deposition activity of ceramicine B, a major constituent of C. ceramicus that showed potent anti-melanin deposition activity, was investigated. Tyrosinase enzymatic activity and tyrosinase mRNA expression were not affected by ceramicine B. The anti-melanin deposition activity of ceramicine B was shown to be related to the downregulation of tyrosinase protein expression. These results suggest that ceramicines have potential to be used as depigmentation agents.

  18. Monitoring Brain Activity with Protein Voltage and Calcium Sensors

    PubMed Central

    Storace, Douglas A.; Braubach, Oliver R.; Jin, Lei; Cohen, Lawrence B.; Sung, Uhna

    2015-01-01

    Understanding the roles of different cell types in the behaviors generated by neural circuits requires protein indicators that report neural activity with high spatio-temporal resolution. Genetically encoded fluorescent protein (FP) voltage sensors, which optically report the electrical activity in distinct cell populations, are, in principle, ideal candidates. Here we demonstrate that the FP voltage sensor ArcLight reports odor-evoked electrical activity in the in vivo mammalian olfactory bulb in single trials using both wide-field and 2-photon imaging. ArcLight resolved fast odorant-responses in individual glomeruli, and distributed odorant responses across a population of glomeruli. Comparisons between ArcLight and the protein calcium sensors GCaMP3 and GCaMP6f revealed that ArcLight had faster temporal kinetics that more clearly distinguished activity elicited by individual odorant inspirations. In contrast, the signals from both GCaMPs were a saturating integral of activity that returned relatively slowly to the baseline. ArcLight enables optical electrophysiology of mammalian neuronal population activity in vivo. PMID:25970202

  19. Gas Diffusion Barriers Using Atomic Layer Deposition: A New Calcium Test and Polymer Substrate Effects

    NASA Astrophysics Data System (ADS)

    Bertrand, Jacob Andrew

    The increasing demand on available energy resources has led to a desire for more energy efficient devices. The wide use of displays in consumer electronics, such as televisions, cell phones, cameras and computers makes them an ideal target for improvement. Organic light-emitting diodes (OLEDs) are a good candidate to replace traditional Si based devices. However, the low work function metals typically used as electrodes in OLEDs are very reactive with water and oxygen. Ultralow permeability gas diffusion barriers with water vapor transmission rates (WVTRs) as low as <10-6g/(m2*day) are required on the polymers used to fabricate organic electronic and thin film photovoltaic devices. Atomic Layer Deposition (ALD) uses self-limiting surface reactions to deposit thin conformal films. ALD is capable of depositing thin, conformal, high quality barriers. WVTR values as low as ˜5 x 10-5 g/(m2*day) have been measured for Al2O3 ALD films at 38 °C/85% RH using the Ca test with optical transmission probing. The Ca test is a technique with very high sensitivity to measure ultralow WVTRs. This test relies on measuring the oxidation of a Ca metal film by monitoring the change in its optical or electrical properties. However, glass lid control experiments have indicated that the WVTRs measured by the Ca test are limited by H2O permeability through the epoxy seals. Varying results have been reported in the literature using the electrical conductance of Ca to measure permeation. In this work, two approaches were applied to overcome the epoxy edge seal limitations. The first approach was to deposit Al2O 3 ALD barriers directly on Ca metal. While the Al 2O3 ALD barriers were successfully deposited, the measurement of an accurate WVTR was limited by barrier pinholes. The presence of pinholes in the Al2O3 ALD barrier on Ca results in the localized oxidation of the Ca sensor. Heterogeneous degradation of the Ca causes inaccuracies in the conductance of the film. As oxidation regions

  20. Calcium influx factor (CIF) as a diffusible messenger for the activation of capacitative calcium entry in Xenopus oocytes.

    PubMed

    Kim, H Y; Hanley, M R

    1999-06-30

    Acid extracts of thapsigargin-treated Xenopus oocytes revealed Ca2(+)-dependent Cl- currents by microinjection into Xenopus oocytes. These currents were detected in highly purified fractions by carrying out a sequence of purification steps including gel filtration chromatography and high performance thin layer chromatography. The nature of the membrane currents evoked by the highly purified fractions were carried by chloride ions as blockade by the selective chloride channel blocker 1 mM niflumic acid. Injection of the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) eradicated the current activities, indicating that the current responses are completely Ca2(+)-dependent. Moreover, the currents were sensitive to the removal of extracellular calcium, indicating the dependence on calcium entry through plasma membrane calcium entry channels. These results elucidate that the highly purified fractions aquired by thapsigargin-stimulated oocytes is an authentic calcium influx factor (CIF). Thus, the detection of increased CIF production from thapsigargin treatment in Xenopus oocytes would give strong support for the existence of CIF as a diffusible messenger for the activation of capacitative calcium entry pathways in Xenopus oocytes.

  1. Synthesis of europium- or terbium-activated calcium tungstate phosphors

    NASA Astrophysics Data System (ADS)

    Forgaciu, Flavia; Popovici, Elisabeth-Jeanne; Ungur, Laura; Vadan, Maria; Vasilescu, Marilena; Nazarov, Mihail

    2001-06-01

    Utilization of luminescent substances in various optoelectronic devices depends on their luminescent properties and sensitivity to various excitation radiation as well as on particle size distribution and crystalline structure of luminous powders. Calcium tungstate phosphors are well excited with roentgen radiation, so that they are largely used for manufacture of x-ray intensifying screens. Being sensitive to short UV-radiation as well, they could be utilized in Plasma Display Panels or in advertising signs fluorescent tubes. In order to diversify the utilization possibilities of this tungstate class, luminescent powders based on CaWO4:Eu3+ and CaWO4:Tb3+ were synthesized and characterized. As compared with the starting self-activated phosphor, larger excitation wavelength domain and emission colors from blue-to-green-to- yellow-to-red were obtained. The good UV excitability and variable luminescence color recommend these phosphors for optoelectronic device manufacture.

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

    PubMed Central

    Hermann, Anton; Sitdikova, Guzel F.; Weiger, Thomas M.

    2015-01-01

    All cells contain ion channels in their outer (plasma) and inner (organelle) membranes. Ion channels, similar to other proteins, are targets of oxidative impact, which modulates ion fluxes across membranes. Subsequently, these ion currents affect electrical excitability, such as action potential discharge (in neurons, muscle, and receptor cells), alteration of the membrane resting potential, synaptic transmission, hormone secretion, muscle contraction or coordination of the cell cycle. In this chapter we summarize effects of oxidative stress and redox mechanisms on some ion channels, in particular on maxi calcium-activated potassium (BK) channels which play an outstanding role in a plethora of physiological and pathophysiological functions in almost all cells and tissues. We first elaborate on some general features of ion channel structure and function and then summarize effects of oxidative alterations of ion channels and their functional consequences. PMID:26287261

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

    PubMed

    Hermann, Anton; Sitdikova, Guzel F; Weiger, Thomas M

    2015-08-17

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

  4. Relationship between structure of phenothiazine analogues and their activity on platelet calcium fluxes.

    PubMed Central

    Enouf, J.; Lévy-Toledano, S.

    1984-01-01

    Phenothiazine analogues have been tested for their effect on calcium uptake into platelet membrane vesicles and on ionophore-induced platelet activation, both phenomena being Ca2+-dependent. Both calcium uptake into membrane vesicles and ionophore-induced platelet activation were inhibited by the drugs. Evidence for two inhibitors as potent as chlorpromazine and trifluoperazine was found. These drugs are apparently competitive inhibitors of calcium uptake. A structure-activity relationship has been established. The data suggest that the phenothiazines are able to inhibit calmodulin-insensitive calcium uptake of platelet membrane vesicles and that therefore they cannot be assumed to be selective inhibitors of calmodulin interactions under all circumstances. PMID:6697061

  5. Antilithiatic Activity of phlorotannin rich extract of Sarghassum Wightii on Calcium Oxalate Urolithiais – In Vitro and In Vivo Evaluation

    PubMed Central

    Sujatha, D.; Singh, Kiranpal; Vohra, Mursalin; Kumar, K. Vijay; Sunitha, S.

    2015-01-01

    ABSTRACT Purpose: Urolithiasis is a common urological disorder responsible for serious human affliction and cost to the society with a high recurrence rate. The aim of the present study was to systematically evaluate the phlorotannin rich extract of Sargassum wightii using suitable in vitro and in vivo models to provide scientific evidence for its antilithiatic activity. Materials and Methods: To explore the effect of Sargassum wightii on calcium oxalate crystallization, in vitro assays like crystal nucleation, aggregation and crystal growth were performed. Calcium oxalate urolithiasis was induced in male Sprague dawley rats using a combination of gentamicin and calculi producing diet (5% ammonium oxalate and rat pellet feed). The biochemical parameters like calcium, oxalate, magnesium, phosphate, sodium and potassium were evaluated in urine, serum and kidney homogenates. Histopathological studies were also done to confirm the biochemical findings. Results: The yield of Sargassum wightii extract was found to be 74.5 gm/kg and confirmed by quantitative analysis. In vitro experiments with Sargassum wightii showed concentration dependent inhibition of calcium oxalate nucleation, aggregation and growth supported by SEM analysis. In the in vivo model, Sargassum wightii reduced both calcium and oxalate supersaturation in urine, serum and deposition in the kidney. The biochemical results were supported by histopathological studies. Conclusion: The findings of the present study suggest that Sargassum wightii has the ability to prevent nucleation, aggregation and growth of calcium oxalate crystals. Sargassum wightii has better preventive effect on calcium oxalate stone formation indicating its strong potential to develop as a therapeutic option to prevent recurrence of urolithiasis. PMID:26200544

  6. Active protein and calcium hydroxyapatite bilayers grown by laser techniques for therapeutic applications.

    PubMed

    Motoc, M M; Axente, E; Popescu, C; Sima, L E; Petrescu, S M; Mihailescu, I N; Gyorgy, E

    2013-09-01

    Active protein and bioceramic calcium hydroxyapatite (HA) bilayers were grown by combining conventional pulsed laser deposition (PLD) and matrix-assisted pulsed laser evaporation (MAPLE) techniques. A pulsed UV KrF* excimer laser was used for the irradiations. The HA layers were grown by PLD. Proteins with antimicrobial action were attached to the bioceramic layers using MAPLE. The composite MAPLE targets were obtained by dissolving the proteins powder in distilled water. The crystalline status and chemical composition of the obtained structures were studied by X-ray diffractometry and Fourier transform infrared spectroscopy. The layers were grown for the design of advanced future metal implants coatings, ensuring both enhanced bone formation and localized antimicrobial therapy. Our results demonstrated that protein coatings improve bone cell proliferation in vitro. Immunofluorescence experiments show that actin filaments stretch throughout bone cells and sustain their optimal spreading.

  7. Continuous cellularization of calcium phosphate hybrid scaffolds induced by plasma polymer activation.

    PubMed

    Bergemann, Claudia; Cornelsen, Matthias; Quade, Antje; Laube, Thorsten; Schnabelrauch, Matthias; Rebl, Henrike; Weißmann, Volker; Seitz, Hermann; Nebe, Barbara

    2016-02-01

    The generation of hybrid materials based on β-tricalcium phosphate (TCP) and various biodegradable polymers like poly(l-lactide-co-d,l-lactide) (PLA) represents a common approach to overcoming the disadvantages of pure TCP devices. These disadvantages lie in TCP's mechanical properties, such as brittleness. The positive characteristic of PLA - improvement of compressive strength of calcium phosphate scaffolds - is diametrically opposed to its cell attractiveness. Therefore, the objective of this work was to optimize osteoblast migration and cellularization inside a three-dimensionally (3D) printed, PLA polymer stabilized TCP hybrid scaffold by a plasma polymer process depositing amino groups via allylamine. MG-63 osteoblastic cells inside the 10mm hybrid scaffold were dynamically cultivated for 14days in a 3D model system integrated in a perfusion reactor. The whole TCP/PLA hybrid scaffold was continuously colonized due to plasma polymerized allylamine activation inducing the migration potential of osteoblasts.

  8. Inorganic lead and calcium interact positively in activation of calmodulin.

    PubMed

    Kern, M; Wisniewski, M; Cabell, L; Audesirk, G

    2000-06-01

    Calmodulin is a ubiquitous calcium-binding protein that mediates many of the intracellular actions of Ca2+ ions. The calcium-binding sites of calmodulin consist of four EF-hand motifs; full activation of calmodulin normally occurs when all four sites are occupied by Ca2+. Inorganic lead (PY2+) has been shown to activate calmodulin at total lead concentrations similar to the concentrations of Ca2+ required for activation (Goldstein and Ar, 1983; Habermann et al., 1983), but the free Pb2+ concentrations required for calmodulin activation have not been determined. In addition, it is possible that activation may occur with different sites occupied by different divalent cations, for example Ca2+ and Pb2+. We investigated the ability of free Pb2+, alone or in combination with Ca2+, to activate calmodulin. In aqueous media, N-phenyl-1-naphthylamine (NPN) and 8-anilino-1-naphthalenesulfonate (ANS) show increased fluorescence when bound to hydrophobic regions of proteins. This increased fluorescence has been used to monitor the conformational change that occurs during calmodulin activation (LaPorte et al., 1980). In the presence of calmodulin, both Ca2+ and Pb2+ stimulated increased fluorescence of NPN and ANS. Threshold and EC50 free metal concentrations were approximately 100 nM and 450-500 nM, respectively, for Ca2+ and 100 pM and 400-550 pM, respectively, for Pb2+. Fluorescence was enhanced by combinations of low concentrations of free Ca2+ and Pb2+; for example, as little as 20 pM free Pb2+ enhanced fluorescence in combination with 200 nM free Ca2+. The activity of the PDE1 isoform of cyclic nucleotide phosphodiesterase is stimulated by Ca2+/calmodulin (Wang et al., 1990). In the presence of calmodulin, we found that Ca2+ and Pb2+ activated calmodulin-stimulated PDE activity, with threshold and EC50 free metal concentrations of approximately 200 nM and 1200 nM, respectively, for Ca2+ and 300 pM and 430 pM, respectively, for Pb2+. PDE activity was stimulated by

  9. A Markovian Entropy Measure for the Analysis of Calcium Activity Time Series.

    PubMed

    Marken, John P; Halleran, Andrew D; Rahman, Atiqur; Odorizzi, Laura; LeFew, Michael C; Golino, Caroline A; Kemper, Peter; Saha, Margaret S

    2016-01-01

    Methods to analyze the dynamics of calcium activity often rely on visually distinguishable features in time series data such as spikes, waves, or oscillations. However, systems such as the developing nervous system display a complex, irregular type of calcium activity which makes the use of such methods less appropriate. Instead, for such systems there exists a class of methods (including information theoretic, power spectral, and fractal analysis approaches) which use more fundamental properties of the time series to analyze the observed calcium dynamics. We present a new analysis method in this class, the Markovian Entropy measure, which is an easily implementable calcium time series analysis method which represents the observed calcium activity as a realization of a Markov Process and describes its dynamics in terms of the level of predictability underlying the transitions between the states of the process. We applied our and other commonly used calcium analysis methods on a dataset from Xenopus laevis neural progenitors which displays irregular calcium activity and a dataset from murine synaptic neurons which displays activity time series that are well-described by visually-distinguishable features. We find that the Markovian Entropy measure is able to distinguish between biologically distinct populations in both datasets, and that it can separate biologically distinct populations to a greater extent than other methods in the dataset exhibiting irregular calcium activity. These results support the benefit of using the Markovian Entropy measure to analyze calcium dynamics, particularly for studies using time series data which do not exhibit easily distinguishable features.

  10. A Markovian Entropy Measure for the Analysis of Calcium Activity Time Series

    PubMed Central

    Rahman, Atiqur; Odorizzi, Laura; LeFew, Michael C.; Golino, Caroline A.; Kemper, Peter; Saha, Margaret S.

    2016-01-01

    Methods to analyze the dynamics of calcium activity often rely on visually distinguishable features in time series data such as spikes, waves, or oscillations. However, systems such as the developing nervous system display a complex, irregular type of calcium activity which makes the use of such methods less appropriate. Instead, for such systems there exists a class of methods (including information theoretic, power spectral, and fractal analysis approaches) which use more fundamental properties of the time series to analyze the observed calcium dynamics. We present a new analysis method in this class, the Markovian Entropy measure, which is an easily implementable calcium time series analysis method which represents the observed calcium activity as a realization of a Markov Process and describes its dynamics in terms of the level of predictability underlying the transitions between the states of the process. We applied our and other commonly used calcium analysis methods on a dataset from Xenopus laevis neural progenitors which displays irregular calcium activity and a dataset from murine synaptic neurons which displays activity time series that are well-described by visually-distinguishable features. We find that the Markovian Entropy measure is able to distinguish between biologically distinct populations in both datasets, and that it can separate biologically distinct populations to a greater extent than other methods in the dataset exhibiting irregular calcium activity. These results support the benefit of using the Markovian Entropy measure to analyze calcium dynamics, particularly for studies using time series data which do not exhibit easily distinguishable features. PMID:27977764

  11. Calcium promotes activity and confers heat stability on plant peroxidases

    PubMed Central

    Plieth, Christoph; Vollbehr, Sonja

    2012-01-01

    In this paper we demonstrate how peroxidase (PO) activities and their heat stability correlate with the availability of free Ca2+ ions. Calcium ions work as a molecular switch for PO activity and exert a protective function, rendering POs heat stable. The concentration ranges of these two activities differ markedly. POs are activated by µM Ca2+ concentration ranges, whereas heat stabilization is observed in the nM range. This suggests the existence of different Ca2+ binding sites. The heat stability of POs depends on the source plant species. Terrestrial plants have POs that exhibit higher temperature stability than those POs from limnic and marine plants. Different POs from a single species can differ in terms of heat stability. The abundance of different POs within a plant is dependent on age and developmental stage. The heat stability of a PO does not necessarily correlate with the maximum temperature the source species is usually exposed to in its natural habitat. This raises questions on the role of POs in the heat tolerance of plants. Consequently, detailed investigations are needed to identify and characterize individual POs, with regard to their genetic origin, subcellular expression, tissue abundance, developmental emergence and their functions in innate and acquired heat tolerance. PMID:22580695

  12. Calcium promotes activity and confers heat stability on plant peroxidases.

    PubMed

    Plieth, Christoph; Vollbehr, Sonja

    2012-06-01

    In this paper we demonstrate how peroxidase (PO) activities and their heat stability correlate with the availability of free Ca(2+) ions. Calcium ions work as a molecular switch for PO activity and exert a protective function, rendering POs heat stable. The concentration ranges of these two activities differ markedly. POs are activated by µM Ca(2+) concentration ranges, whereas heat stabilization is observed in the nM range. This suggests the existence of different Ca(2+) binding sites. The heat stability of POs depends on the source plant species. Terrestrial plants have POs that exhibit higher temperature stability than those POs from limnic and marine plants. Different POs from a single species can differ in terms of heat stability. The abundance of different POs within a plant is dependent on age and developmental stage. The heat stability of a PO does not necessarily correlate with the maximum temperature the source species is usually exposed to in its natural habitat. This raises questions on the role of POs in the heat tolerance of plants. Consequently, detailed investigations are needed to identify and characterize individual POs, with regard to their genetic origin, subcellular expression, tissue abundance, developmental emergence and their functions in innate and acquired heat tolerance.

  13. Intracellular calcium strongly potentiates agonist-activated TRPC5 channels

    PubMed Central

    Blair, Nathaniel T.; Kaczmarek, J. Stefan

    2009-01-01

    TRPC5 is a calcium (Ca2+)-permeable nonselective cation channel expressed in several brain regions, including the hippocampus, cerebellum, and amygdala. Although TRPC5 is activated by receptors coupled to phospholipase C, the precise signaling pathway and modulatory signals remain poorly defined. We find that during continuous agonist activation, heterologously expressed TRPC5 currents are potentiated in a voltage-dependent manner (∼5-fold at positive potentials and ∼25-fold at negative potentials). The reversal potential, doubly rectifying current–voltage relation, and permeability to large cations such as N-methyl-d-glucamine remain unchanged during this potentiation. The TRPC5 current potentiation depends on extracellular Ca2+: replacement by Ba2+ or Mg2+ abolishes it, whereas the addition of 10 mM Ca2+ accelerates it. The site of action for Ca2+ is intracellular, as simultaneous fura-2 imaging and patch clamp recordings indicate that potentiation is triggered at ∼1 µM [Ca2+]. This potentiation is prevented when intracellular Ca2+ is tightly buffered, but it is promoted when recording with internal solutions containing elevated [Ca2+]. In cell-attached and excised inside-out single-channel recordings, increases in internal [Ca2+] led to an ∼10–20-fold increase in channel open probability, whereas single-channel conductance was unchanged. Ca2+-dependent potentiation should result in TRPC5 channel activation preferentially during periods of repetitive firing or coincident neurotransmitter receptor activation. PMID:19398778

  14. Nutritional impact of elevated calcium transport activity in carrots

    PubMed Central

    Morris, Jay; Hawthorne, Keli M.; Hotze, Tim; Abrams, Steven A.; Hirschi, Kendal D.

    2008-01-01

    Nutrition recommendations worldwide emphasize ingestion of plant-based diets rather than diets that rely primarily on animal products. However, this plant-based diet could limit the intake of essential nutrients such as calcium. Osteoporosis is one of the world's most prevalent nutritional disorders, and inadequate dietary calcium is a known contributor to the pathophysiology of this condition. Previously, we have modified carrots to express increased levels of a plant calcium transporter (sCAX1), and these plants contain ≈2-fold-higher calcium content in the edible portions of the carrots. However, it was unproven whether this change would increase the total amount of bioavailable calcium. In randomized trials, we labeled these modified carrots with isotopic calcium and fed them to mice and humans to assess calcium bioavailability. In mice feeding regimes (n = 120), we measured 45Ca incorporation into bones and determined that mice required twice the serving size of control carrots to obtain the calcium found in sCAX1 carrots. We used a dual-stable isotope method with 42Ca-labeled carrots and i.v. 46Ca to determine the absorption of calcium from these carrots in humans. In a cross-over study of 15 male and 15 female adults, we found that when people were fed sCAX1 and control carrots, total calcium absorption per 100 g of carrots was 41% ± 2% higher in sCAX1 carrots. Both the mice and human feeding studies demonstrate increased calcium absorption from sCAX1-expressing carrots compared with controls. These results demonstrate an alternative means of fortifying vegetables with bioavailable calcium. PMID:18202180

  15. Activation of PAC1 Receptors in Rat Cerebellar Granule Cells Stimulates Both Calcium Mobilization from Intracellular Stores and Calcium Influx through N-Type Calcium Channels

    PubMed Central

    Basille-Dugay, Magali; Vaudry, Hubert; Fournier, Alain; Gonzalez, Bruno; Vaudry, David

    2013-01-01

    High concentrations of pituitary adenylate cyclase-activating polypeptide (PACAP) and a high density of PACAP binding sites have been detected in the developing rat cerebellum. In particular, PACAP receptors are actively expressed in immature granule cells, where they activate both adenylyl cyclase and phospholipase C. The aim of the present study was to investigate the ability of PACAP to induce calcium mobilization in cerebellar granule neurons. Administration of PACAP-induced a transient, rapid, and monophasic rise of the cytosolic calcium concentration ([Ca2+]i), while vasoactive intestinal peptide was devoid of effect, indicating the involvement of the PAC1 receptor in the Ca2+ response. Preincubation of granule cells with the Ca2+ ATPase inhibitor, thapsigargin, or the d-myo-inositol 1,4,5-trisphosphate (IP3) receptor antagonist, 2-aminoethoxydiphenyl borate, markedly reduced the stimulatory effect of PACAP on [Ca2+]i. Furthermore, addition of the calcium chelator, EGTA, or exposure of cells to the non-selective Ca2+ channel blocker, NiCl2, significantly attenuated the PACAP-evoked [Ca2+]i increase. Preincubation of granule neurons with the N-type Ca2+ channel blocker, ω-conotoxin GVIA, decreased the PACAP-induced [Ca2+]i response, whereas the L-type Ca2+ channel blocker, nifedipine, and the P- and Q-type Ca2+ channel blocker, ω-conotoxin MVIIC, had no effect. Altogether, these findings indicate that PACAP, acting through PAC1 receptors, provokes an increase in [Ca2+]i in granule neurons, which is mediated by both mobilization of calcium from IP3-sensitive intracellular stores and activation of N-type Ca2+ channel. Some of the activities of PACAP on proliferation, survival, migration, and differentiation of cerebellar granule cells could thus be mediated, at least in part, through these intracellular and/or extracellular calcium fluxes. PMID:23675369

  16. Phase Transformation and Morphology of Calcium Phosphate Prepared by Electrochemical Deposition Process Through Alkali Treatment and Calcination

    NASA Astrophysics Data System (ADS)

    Chen, Hui-Ting; Wang, Moo-Chin; Chang, Kuo-Ming; Wang, Szu-Hao; Shih, Wei-Jen; Li, Wong-Long

    2014-04-01

    The phase transformation and morphology of calcium phosphate prepared by the electrochemical deposition (ECD) process through alkali treatment and calcination have been characterized using X-ray diffraction (XRD), thermogravimetry and differential thermal analyses (TG/DTA), and scanning electron microscopy (SEM). At the ECD process, when the excess OH- was produced, the reaction of 10Ca2++6PO{4/3-}+2OH-→Ca10(PO4)6(OH)2 takes place on the Ti-6Al-4V and the HA is deposited. The XRD results reveal that the as-deposit was mostly composed of dicalcium phosphate dehydrate (Ca2H4P2O9; DCPD) and the minor phase of hydroxyapatite (Ca10(PO4)6(OH)2; HA). After NaOH treatment, all DCPD were converted to HA. Moreover, the content of HA phase increases with ECD potential. After being calcined at 673 K and 873 K (400 °C and 600 °C) for 4 hours, the phase of HA maintained the major phase for an alkali-treated deposited sample. After being calcined at 1073 K (800 °C) for 4 hours, some HA decomposed and caused the minor phases of β-tricalcium phosphate ( β-Ca3(PO4)2; β-TCP), calcium pyrophosphate (Ca2P2O7; CPP), and calcium oxide (CaO) formation. The β-TCP becomes the major phase with residual HA and CaO after being calcined at 1273 K (1000 °C) for 4 hours. The crack forms due to the release of absorbed water from the interior to top surface of sample. For the as-alkali treatment samples, the microstructures were affected by ECD potentials; when the deposited samples after alkali treatment and calcined at 1073 K (800 °C) for 4 hours, the microstructure presents the need-like "preforming HA" (pre-HA) from the matrix of plate-like postforming HA (post-HA).

  17. Calcium oxalate crystal deposition in a patient with Aspergilloma due to Aspergillus niger

    PubMed Central

    Oda, Miku; Wakayama, Megumi; Shibuya, Kazutoshi; Ogawa, Yukari; Inui, Toshiya; Yokoyama, Emi; Inoue, Manami; Shimoyamada, Hiroaki; Fujiwara, Masachika; Ota, Tomohiro; Takizawa, Hajime; Goto, Hajime

    2013-01-01

    Discrimination between aspergilloma and chronic necrotizing pulmonary aspergillosis (CNPA) based on radiological findings can difficult. We describe a patient with aspergilloma and organizing pneumonia that was possibly caused by Aspergillus niger infection and radiologically mimicked CNPA. A postmortem histological analysis showed diffuse alveolar damage that had originated in peri-cavitary lung parenchyma. Calcium oxalate or Aspergillus niger was located inside, but not outside the cavity in the right upper lobe. Calcium oxalate or other unknown hyphal bioactive components might provoke severe lung inflammation not only adjacent to the cavity, but also on the contralateral side. PMID:23991333

  18. Potentiostatic pulse-deposition of calcium phosphate on magnesium alloy for temporary implant applications--an in vitro corrosion study.

    PubMed

    Kannan, M Bobby; Wallipa, O

    2013-03-01

    In this study, a magnesium alloy (AZ91) was coated with calcium phosphate using potentiostatic pulse-potential and constant-potential methods and the in vitro corrosion behaviour of the coated samples was compared with the bare metal. In vitro corrosion studies were carried out using electrochemical impedance spectroscopy and potentiodynamic polarization in simulated body fluid (SBF) at 37 °C. Calcium phosphate coatings enhanced the corrosion resistance of the alloy, however, the pulse-potential coating performed better than the constant-potential coating. The pulse-potential coating exhibited ~3 times higher polarization resistance than that of the constant-potential coating. The corrosion current density obtained from the potentiodynamic polarization curves was significantly less (~60%) for the pulse-deposition coating as compared to the constant-potential coating. Post-corrosion analysis revealed only slight corrosion on the pulse-potential coating, whereas the constant-potential coating exhibited a large number of corrosion particles attached to the coating. The better in vitro corrosion performance of the pulse-potential coating can be attributed to the closely packed calcium phosphate particles.

  19. Allosteric activation of apicomplexan calcium-dependent protein kinases

    PubMed Central

    Ingram, Jessica R.; Knockenhauer, Kevin E.; Markus, Benedikt M.; Mandelbaum, Joseph; Ramek, Alexander; Shan, Yibing; Shaw, David E.; Schwartz, Thomas U.; Ploegh, Hidde L.; Lourido, Sebastian

    2015-01-01

    Calcium-dependent protein kinases (CDPKs) comprise the major group of Ca2+-regulated kinases in plants and protists. It has long been assumed that CDPKs are activated, like other Ca2+-regulated kinases, by derepression of the kinase domain (KD). However, we found that removal of the autoinhibitory domain from Toxoplasma gondii CDPK1 is not sufficient for kinase activation. From a library of heavy chain-only antibody fragments (VHHs), we isolated an antibody (1B7) that binds TgCDPK1 in a conformation-dependent manner and potently inhibits it. We uncovered the molecular basis for this inhibition by solving the crystal structure of the complex and simulating, through molecular dynamics, the effects of 1B7–kinase interactions. In contrast to other Ca2+-regulated kinases, the regulatory domain of TgCDPK1 plays a dual role, inhibiting or activating the kinase in response to changes in Ca2+ concentrations. We propose that the regulatory domain of TgCDPK1 acts as a molecular splint to stabilize the otherwise inactive KD. This dependence on allosteric stabilization reveals a novel susceptibility in this important class of parasite enzymes. PMID:26305940

  20. Allosteric activation of apicomplexan calcium-dependent protein kinases

    SciTech Connect

    Ingram, Jessica R.; Knockenhauer, Kevin E.; Markus, Benedikt M.; Mandelbaum, Joseph; Ramek, Alexander; Shan, Yibing; Shaw, David E.; Schwartz, Thomas U.; Ploegh, Hidde L.; Lourido, Sebastian

    2015-08-24

    Calcium-dependent protein kinases (CDPKs) comprise the major group of Ca2+-regulated kinases in plants and protists. It has long been assumed that CDPKs are activated, like other Ca2+-regulated kinases, by derepression of the kinase domain (KD). However, we found that removal of the autoinhibitory domain from Toxoplasma gondii CDPK1 is not sufficient for kinase activation. From a library of heavy chain-only antibody fragments (VHHs), we isolated an antibody (1B7) that binds TgCDPK1 in a conformation-dependent manner and potently inhibits it. We uncovered the molecular basis for this inhibition by solving the crystal structure of the complex and simulating, through molecular dynamics, the effects of 1B7–kinase interactions. In contrast to other Ca2+-regulated kinases, the regulatory domain of TgCDPK1 plays a dual role, inhibiting or activating the kinase in response to changes in Ca2+ concentrations. We propose that the regulatory domain of TgCDPK1 acts as a molecular splint to stabilize the otherwise inactive KD. This dependence on allosteric stabilization reveals a novel susceptibility in this important class of parasite enzymes.

  1. Allosteric activation of apicomplexan calcium-dependent protein kinases

    DOE PAGES

    Ingram, Jessica R.; Knockenhauer, Kevin E.; Markus, Benedikt M.; ...

    2015-08-24

    Calcium-dependent protein kinases (CDPKs) comprise the major group of Ca2+-regulated kinases in plants and protists. It has long been assumed that CDPKs are activated, like other Ca2+-regulated kinases, by derepression of the kinase domain (KD). However, we found that removal of the autoinhibitory domain from Toxoplasma gondii CDPK1 is not sufficient for kinase activation. From a library of heavy chain-only antibody fragments (VHHs), we isolated an antibody (1B7) that binds TgCDPK1 in a conformation-dependent manner and potently inhibits it. We uncovered the molecular basis for this inhibition by solving the crystal structure of the complex and simulating, through molecular dynamics,more » the effects of 1B7–kinase interactions. In contrast to other Ca2+-regulated kinases, the regulatory domain of TgCDPK1 plays a dual role, inhibiting or activating the kinase in response to changes in Ca2+ concentrations. We propose that the regulatory domain of TgCDPK1 acts as a molecular splint to stabilize the otherwise inactive KD. This dependence on allosteric stabilization reveals a novel susceptibility in this important class of parasite enzymes.« less

  2. Composition and source identification of deposits forming in landfill gas (LFG) engines and effect of activated carbon treatment on deposit composition.

    PubMed

    Sevimoğlu, Orhan; Tansel, Berrin

    2013-10-15

    Compositions of deposits forming on engines parts operated with landfill gas (LFG) were analyzed. The deposit compositions were compared before and after the installation of activated carbon system for treatment of LFG. Deposits forming on the spark plugs had significantly higher levels of calcium, chromium, and nickel in comparison to those forming on the engine heads. The LFG contained about 9.5 ± 0.4 mg/m(3) total siloxanes, majority of which were octamethylcyclotetrasiloxane (D4) (5.0 ± 0.2 mg/m(3)), decamethylcyclopentasiloxane (D5) (2.9 ± 0.1 mg/m(3)) and hexamethyldisiloxane (L2) (1.6 ± 0.1 mg/m(3)). The samples collected from the engine heads before the activated carbon treatment of LFG had significantly high levels of silicon (149,400 ± 89,900 mg/kg) as well as calcium (70,840 ± 17,750 mg/kg), sulfur (42,500 ± 11,500 mg/kg), and zinc (22,300 ± 7200 mg/kg). After the activated carbon treatment, silicon levels decreased significantly; however, deposits had higher sulfur content (104,560 ± 68,100 mg/kg) indicating that the activated carbon released some sulfur during treatment. The analyses indicate that zinc and calcium originated from the additives in the lube oil while lead, aluminum, copper, nickel, iron, chromium were due to the engine wear.

  3. Calcium Deposits in the Crayfish, Cherax quadricarinatus: Microstructure Versus Elemental Distribution.

    PubMed

    Luquet, Gilles; Dauphin, Yannicke; Percot, Aline; Salomé, Murielle; Ziegler, Andreas; Fernández, Maria S; Arias, José L

    2016-02-01

    The crayfish Cherax quadricarinatus stores calcium ions, easily mobilizable after molting, for calcifying parts of the new exoskeleton. They are chiefly stored as amorphous calcium carbonate (ACC) during each premolt in a pair of gastroliths synthesized in the stomach wall. How calcium carbonate is stabilized in the amorphous state in such a biocomposite remains speculative. The knowledge of the microstructure at the nanometer level obtained by field emission scanning electron microscopy and atomic force microscopy combined with scanning electron microscopy energy-dispersive X-ray spectroscopy, micro-Raman and X-ray absorption near edge structure spectroscopy gave relevant information on the elaboration of such an ACC-stabilized biomineral. We observed nanogranules distributed along chitin-protein fibers and the aggregation of granules in thin layers. AFM confirmed the nanolevel structure, showing granules probably surrounded by an organic layer and also revealing a second level of aggregation as described for other crystalline biominerals. Raman analyses showed the presence of ACC, amorphous calcium phosphate, and calcite. Elemental analyses confirmed the presence of elements like Fe, Na, Mg, P, and S. P and S are heterogeneously distributed. P is present in both the mineral and organic phases of gastroliths. S seems present as sulfate (probably as sulfated sugars), sulfonate, sulfite, and sulfoxide groups and, in a lesser extent, as sulfur-containing amino acids.

  4. Reconstruction of burst activity from calcium imaging of neuronal population via Lq minimization and interval screening

    PubMed Central

    Quan, Tingwei; Lv, Xiaohua; Liu, Xiuli; Zeng, Shaoqun

    2016-01-01

    Calcium imaging is becoming an increasingly popular technology to indirectly measure activity patterns in local neuronal networks. Based on the dependence of calcium fluorescence on neuronal spiking, two-photon calcium imaging affords single-cell resolution of neuronal population activity. However, it is still difficult to reconstruct neuronal activity from complex calcium fluorescence traces, particularly for traces contaminated by noise. Here, we describe a robust and efficient neuronal-activity reconstruction method that utilizes Lq minimization and interval screening (IS), which we refer to as LqIS. The simulation results show that LqIS performs satisfactorily in terms of both accuracy and speed of reconstruction. Reconstruction of simulation and experimental data also shows that LqIS has advantages in terms of the recall rate, precision rate, and timing error. Finally, LqIS is demonstrated to effectively reconstruct neuronal burst activity from calcium fluorescence traces recorded from large-size neuronal population. PMID:27375930

  5. Cellular Mechanisms of Calcium-Mediated Triggered Activity

    NASA Astrophysics Data System (ADS)

    Song, Zhen

    Life-threatening cardiac arrhythmias continue to pose a major health problem. Ventricular fibrillation, which is a complex form of electrical wave turbulence in the lower chambers of the heart, stops the heart from pumping and is the largest cause of natural death in the United States. Atrial fibrillation, a related form of wave turbulence in the upper heart chambers, is in turn the most common arrhythmia diagnosed in clinical practice. Despite extensive research to date, mechanisms of cardiac arrhythmias remain poorly understood. It is well established that both spatial disorder of the refractory period of heart cells and triggered activity (TA) jointly contribute to the initiation and maintenance of arrhythmias. TA broadly refers to the abnormal generation of a single or a sequence of abnormal excitation waves from a small submillimeter region of the heart in the interval of time between two normal waves generated by the heart's natural pacemaker (the sinoatrial node). TA has been widely investigated experimentally and occurs in several pathological conditions where the intracellular concentration of free Ca2+ ions in heart cells becomes elevated. Under such conditions, Ca2+ can be spontaneously released from intracellular stores, thereby driving an electrogenic current that exchanges 3Na+ ions for one Ca2+ ion across the cell membrane. This current in turn depolarizes the membrane of heart cells after a normal excitation. If this calcium-mediated "delayed after depolarization'' (DAD) is sufficiently large, it can generate an action potential. While the arrhythmogenic importance of spontaneous Ca2+ release and DADs is well appreciated, the conditions under which they occur in heart pathologies remain poorly understood. Calcium overload is only one factor among several other factors that can promote DADs, including sympathetic nerve stimulation, different expression levels of membrane ion channels and calcium handling proteins, and different mutations of those

  6. Promotion of beta-glucan synthase activity in corn microsomal membranes by calcium and protein phosphorylation

    NASA Technical Reports Server (NTRS)

    Paliyath, G.; Poovaiah, B. W.

    1988-01-01

    Regulation of the activity of beta-glucan synthase was studied using microsomal preparations from corn coleoptiles. The specific activity as measured by the incorporation of glucose from uridine diphospho-D-[U-14C]glucose varied between 5 to 15 pmol (mg protein)-1 min-1. Calcium promoted beta-glucan synthase activity and the promotion was observed at free calcium concentrations as low as 1 micromole. Kinetic analysis of substrate-velocity curve showed an apparent Km of 1.92 x 10(-4) M for UDPG. Calcium increased the Vmax from 5.88 x 10(-7) mol liter-1 min-1 in the absence of calcium to 9.52 x 10(-7) mol liter-1 min-1 and 1.66 x 10(-6) mol liter-1 min-1 in the presence of 0.5 mM and 1 mM calcium, respectively. The Km values remained the same under these conditions. Addition of ATP further increased the activity above the calcium-promoted level. Sodium fluoride, a phosphoprotein phosphatase inhibitor, promoted glucan synthase activity indicating that phosphorylation and dephosphorylation are involved in the regulation of the enzyme activity. Increasing the concentration of sodium fluoride from 0.25 mM to 10 mM increased glucan synthase activity five-fold over the + calcium + ATP control. Phosphorylation of membrane proteins also showed a similar increase under these conditions. Calmodulin, in the presence of calcium and ATP stimulated glucan synthase activity substantially, indicating that calmodulin could be involved in the calcium-dependent phosphorylation and promotion of beta-glucan synthase activity. The role of calcium in mediating auxin action is discussed.

  7. Deposition of calcium phosphate coatings using condensed phosphates (P2O7(4-) and P3O10(5-)) as phosphate source through induction heating.

    PubMed

    Zhou, Huan; Hou, Saisai; Zhang, Mingjie; Yang, Mengmeng; Deng, Linhong; Xiong, Xinbo; Ni, Xinye

    2016-12-01

    In present work condensed phosphates (P2O7(4-) and P3O10(5-)) were used as phosphate source in induction heating to deposit calcium phosphate coatings. The phase, morphology, and composition of different phosphate-related coatings were characterized and compared using XRD, FTIR, and SEM analyses. Results showed that P2O7(4-)formed calcium pyrophosphate hydrate coatings with interconnected cuboid-like particles. The as-deposited calcium tripolyphosphate hydrate coating with P3O10(5-) was mainly composed of flower-like particles assembled by plate-like crystals. The bioactivity and cytocompatibility of the coatings were also studied. Moreover, the feasibility of using hybrid phosphate sources for preparing and depositing coatings onto magnesium alloy was investigated.

  8. Platelet activating factor raises intracellular calcium ion concentration in macrophages

    PubMed Central

    1986-01-01

    Peritoneal cells from thioglycollate-stimulated mice were allowed to adhere to coverglasses for 2 h to give a dense monolayer of adherent cells greater than 95% of which were macrophages. After incubation with the tetra-acetoxymethyl ester of quin2, coverglasses were rinsed with Ca2+-free saline, oriented at a 45 degree angle in square cuvettes containing a magnetically driven stir bar, and analyzed for changes in quin2 fluorescence in a spectrofluorimeter. Such fluorescence, taken as an indication of intracellular calcium ion concentration ([Ca2+]i), increased as exogenous calcium ion concentration ([Ca2+]o) was raised to 1 mM. At [Ca2+]o approximately equal to 10 microM, [Ca2+]i = 72 +/- 14 nM (n = 26); at [Ca2+]o = 1 mM, [Ca2+]i = 140-220 nM, levels not increased by N, N, N', N'-tetrakis (2-pyridylmethyl) ethylenediamine, a membrane-permeant chelator of heavy metals than can quench quin2. Addition of mouse alpha + beta fibroblast interferon, lipopolysaccharide, thrombin, collagen, vasopressin, ADP, compound 48/80, or U46619 did not change [Ca2+]i. However, addition of platelet activating factor (PAF) (2-20 ng/ml) raised [Ca2+]i by 480 nM within 1 min if [Ca2+]o = 1 mM. In the presence of 5 mM EGTA, PAF raised [Ca2+]i by 25 nM. This suggests that PAF causes influx of exogenous Ca2+, as well as releasing some Ca2+ from intracellular stores. Consistent with these results, when PAF was added to 1 mM Ca2+ in the presence of 100 microM Cd2+ or Mn2+ to block Ca2+ influx, [Ca2+]i increased by only intermediate amounts; at the times of such dampened peak response, [Ca2+]i could be raised within 1 min to normal PAF-stimulated levels by chelation of the exogenous heavy metals with diethylenetriaminepentaacetic acid. Normal PAF responses were observed in the presence of indomethacin. The lowest dose of PAF observed to raise [Ca2+]i was 0.1 ng/ml. Response of [Ca2+]i to 2-20 ng/ml PAF was transient, and second applications had no effect. The PAF response also was seen in

  9. Zinc modulation of calcium activity at the photoreceptor terminal: a calcium imaging study.

    PubMed

    Anastassov, Ivan; Shen, Wen; Ripps, Harris; Chappell, Richard L

    2013-07-01

    There is abundant experimental evidence that zinc ions (Zn(2+)) are present in the synaptic vesicles of vertebrate photoreceptors, and that they are co-released with glutamate. Here we show that increasing the concentration of extracellular zinc (2 μM-2 mM) suppresses the entry of calcium into the synaptic terminals of isolated salamander double cones. The resultant dose-dependent curve was fit by an inverse Hill equation having an IC50 of 38 μM, and Hill coefficient of 1.1. Because there is currently no reliable way to measure the concentration of extracellular zinc, it is not known whether the zinc released under normal circumstances is of physiological significance. In an attempt to circumvent this problem we used zinc chelators to reduce the available pool of endogenous zinc. This enabled us to determine how the absence of zinc affected calcium entry. We found that when intra- or extra-cellular zinc was chelated by 250 μM of membrane-permeable TPEN or 500 μM of membrane-impermeable histidine, there was a significant rise in the depolarization-induced intracellular calcium level within photoreceptor terminals. This increase in internal [Ca(2+)] will undoubtedly lead to a concomitant increase in glutamate release. In addition, we found that blocking the L-type calcium channels that are expressed on the synaptic terminals of photoreceptors with 50 μM nicardipine or 100 μM verapamil abolished the effects of zinc chelation. These findings are a good indication that, when released in vivo, the zinc concentration is sufficient to suppress voltage-gated calcium channels, and reduce the rate of glutamate release from photoreceptor terminals.

  10. Calcium-activated conductance in skate electroreceptors: current clamp experiments.

    PubMed

    Clusin, W T; Bennett, M V

    1977-02-01

    When current clamped, skate electroreceptor epithelium produces large action potentials in response to stimuli that depolarize the lumenal faces of the receptor cells. With increasing stimulus strength these action potentials become prolonged. When the peak voltage exceeds about 140 mV the repolarizing phase is blocked until the end of the stimulus. Perfusion experiments show that the rising phase of the action potential results from an increase in calcium permeability in the lumenal membranes. Perfusion of the lumen with cobalt or with a zero calcium solution containing EGTA blocks the action potential. Perfusion of the lumen with a solution containing 10 mM Ca and 20 mM EGTA initially slows the repolarizing process at all voltages and lowers the potential at which it is blocked. With prolonged perfusion, repolarization is blocked at all voltages. When excitability is abolished by perfusion with cobalt, or with a zero calcium solution containing EGTA, no delayed rectification occurs. We suggest that repolarization during the action potential depends on an influx of calcium into the cytoplasm, and that the rate of repolarization depends on the magnitude of the inward calcium current. Increasingly large stimuli reduce the rate of repolarization by reducing the driving force for calcium, and then block repolarization by causing the lumenal membrane potential to exceed ECa. Changes in extracellular calcium affect repolarization in a manner consistent with the resulting change in ECa.

  11. Hippocalcin gates the calcium activation of the slow afterhyperpolarization in hippocampal pyramidal cells.

    PubMed

    Tzingounis, Anastassios V; Kobayashi, Masaaki; Takamatsu, Ken; Nicoll, Roger A

    2007-02-15

    In the brain, calcium influx following a train of action potentials activates potassium channels that mediate a slow afterhyperpolarization current (I(sAHP)). The key steps between calcium influx and potassium channel activation remain unknown. Here we report that the key intermediate between calcium and the sAHP channels is the diffusible calcium sensor hippocalcin. Brief depolarizations sufficient to activate the I(sAHP) in wild-type mice do not elicit the I(sAHP) in hippocalcin knockout mice. Introduction of hippocalcin in cultured hippocampal neurons leads to a pronounced I(sAHP), while neurons expressing a hippocalcin mutant lacking N-terminal myristoylation exhibit a small I(sAHP) that is similar to that recorded in uninfected neurons. This implies that hippocalcin must bind to the plasma membrane to mediate its effects. These findings support a model in which the calcium sensor for the sAHP channels is not preassociated with the channel complex.

  12. Oxygen nano-bubble water reduces calcium oxalate deposits and tubular cell injury in ethylene glycol-treated rat kidney.

    PubMed

    Hirose, Yasuhiko; Yasui, Takahiro; Taguchi, Kazumi; Fujii, Yasuhiro; Niimi, Kazuhiro; Hamamoto, Shuzo; Okada, Atsushi; Kubota, Yasue; Kawai, Noriyasu; Itoh, Yasunori; Tozawa, Keiichi; Sasaki, Shoichi; Kohri, Kenjiro

    2013-08-01

    Renal tubular cell injury induced by oxalate plays an important role in kidney stone formation. Water containing oxygen nano-bubbles (nanometer-sized bubbles generated from oxygen micro-bubbles; ONB) has anti-inflammatory effects. Therefore, we investigated the inhibitory effects of ONB water on kidney stone formation in ethylene glycol (EG)-treated rats. We divided 60 rats, aged 4 weeks, into 5 groups: control, the water-fed group; 100 % ONB, the 100 % ONB water-fed group; EG, the EG treated water-fed group; EG + 50 % ONB and EG + 100 % ONB, water containing EG and 50 % or 100 % ONB, respectively. Renal calcium oxalate (CaOx) deposition, urinary excretion of N-acetyl-β-D-glucosaminidase (NAG), and renal expression of inflammation-related proteins, oxidative stress biomarkers, and the crystal-binding molecule hyaluronic acid were compared among the 5 groups. In the control and 100 % ONB groups, no renal CaOx deposits were detected. In the EG + 50 % ONB and EG + 100 % ONB groups, ONB water significantly decreased renal CaOx deposits, urinary NAG excretion, and renal monocyte chemoattractant protein-1, osteopontin, and hyaluronic acid expression and increased renal superoxide dismutase-1 expression compared with the EG group. ONB water substantially affected kidney stone formation in the rat kidney by reducing renal tubular cell injury. ONB water is a potential prophylactic agent for kidney stones.

  13. The role of oxygen in the deposition of copper-calcium thin film as diffusion barrier for copper metallization

    NASA Astrophysics Data System (ADS)

    Yu, Zhinong; Ren, Ruihuang; Xue, Jianshe; Yao, Qi; Li, Zhengliang; Hui, Guanbao; Xue, Wei

    2015-02-01

    The properties of copper (Cu) metallization based on copper-calcium (CuCa) diffusion barrier as a function of oxygen flux in the CuCa film deposition were investigated in view of adhesion, diffusion and electronic properties. The CuCa film as the diffusion barrier of Cu film improves the adhesion of Cu film, however, and increases the resistance of Cu film. The introduction of oxygen into the deposition of CuCa film induces the improvement of adhesion and crystallinity of Cu film, but produces a slight increase of resistance. The increased resistance results from the partial oxidation of Cu film. The annealing process in vacuum further improves the adhesion, crystallinity and conductivity of Cu film. X-ray diffraction (XRD) and Auger electron spectroscopy (AES) show that the CuCa alloy barrier layer deposited at oxygen atmosphere has perfect anti-diffusion between Cu film and substrate due to the formation of Ca oxide in the interface of CuCa/substrate.

  14. [Microbial geochemical calcium cycle].

    PubMed

    Zavarzin, G A

    2002-01-01

    The participation of microorganisms in the geochemical calcium cycle is the most important factor maintaining neutral conditions on the Earth. This cycle has profound influence on the fate of inorganic carbon, and, thereby, on the removal of CO2 from the atmosphere. The major part of calcium deposits was formed in the Precambrian, when prokaryotic biosphere predominated. After that, calcium recycling based on biogenic deposition by skeletal organisms became the main process. Among prokaryotes, only a few representatives, e.g., cyanobacteria, exhibit a special calcium function. The geochemical calcium cycle is made possible by the universal features of bacteria involved in biologically mediated reactions and is determined by the activities of microbial communities. In the prokaryotic system, the calcium cycle begins with the leaching of igneous rock predominantly through the action of the community of organotrophic organisms. The release of carbon dioxide to the soil air by organotrophic aerobes leads to leaching with carbonic acid and soda salinization. Under anoxic conditions, of major importance is the organic acid production by primary anaerobes (fermentative microorganisms). Calcium carbonate is precipitated by secondary anaerobes (sulfate reducers) and to a smaller degree by methanogens. The role of the cyanobacterial community in carbonate deposition is exposed by stromatolites, which are the most common organo-sedimentary Precambrian structures. Deposition of carbonates in cyanobacterial mats as a consequence of photoassimilation of CO2 does not appear to be a significant process. It is argued that carbonates were deposited at the boundary between the "soda continent", which emerged as a result of subaerial leaching with carbonic acid, and the ocean containing Ca2+. Such ecotones provided favorable conditions for the development of the benthic cyanobacterial community, which was a precursor of stromatolites.

  15. Relations between coke deposition and activity of HDS catalysts

    SciTech Connect

    Brito, J.; Golding, R.; Severino, F.; Laine, J.

    1982-09-01

    Results of studies of coke deposition due to degradation of 1,3-butadiene at 400/sup 0/C are reported for studies employing supported molybdate catalysts, with and without promoters (Co and Ni) and with or without presulfiding. Initial hydrosulfurization (HDS) behavior of the catalysts was also examined. The results suggest that deposition of coke is one of the reasons for the difference in catalyst activity, and higher initial and steady state activities of presulfided catalysts suggest that H/sub 2/S treatment reduces the deactivation processes such as coke deposition. The cobalt promoted catalysts were found to be more prone to coke formation that the nickel promoted catalysts. (BLM)

  16. Influence of pulse ratio on codeposition of copper species with calcium phosphate coatings on titanium by means of electrochemically assisted deposition.

    PubMed

    Wolf-Brandstetter, Cornelia; Oswald, Steffen; Bierbaum, Susanne; Wiesmann, Hans-Peter; Scharnweber, Dieter

    2014-01-01

    Aim of this study was to combine the well-known biocompatibility and ostoeconductivity of thin calcium phosphate coatings on titanium with proangiogenic signals from codeposited copper species. Copper species could be integrated in mineral layers based on hydroxyapatite by means of electrochemically assisted deposition from electrolytes containing calcium, phosphate, and copper ions. Different combinations of duration and intensity of galvanostatic pulses result in different amounts of deposited calcium phosphate and of copper species even for the same applied total charge. Absolute amounts of copper varied between 2.1 and 6.9 μg/cm², and the copper was distributed homogeneously as shown by EDX mapping. The presence of copper did not change the crystalline phase of deposited calcium phosphate (hydroxyapatite) but provoked a significant decrease in deposited amounts by factor 3 to 4. The copper was deposited mainly as Cu(I) species with a minor fraction of basic copper phosphates. Reduction of copper occurred not only at the surface of titanium but also within the hydroxyapatite coating due to the reaction with hydrogen produced by the electrolysis of water during the cathodic polarization of the substrate.

  17. Calcium-activated chloride channels in the apical region of mouse vomeronasal sensory neurons

    PubMed Central

    Dibattista, Michele; Amjad, Asma; Maurya, Devendra Kumar; Sagheddu, Claudia; Montani, Giorgia; Tirindelli, Roberto

    2012-01-01

    The rodent vomeronasal organ plays a crucial role in several social behaviors. Detection of pheromones or other emitted signaling molecules occurs in the dendritic microvilli of vomeronasal sensory neurons, where the binding of molecules to vomeronasal receptors leads to the influx of sodium and calcium ions mainly through the transient receptor potential canonical 2 (TRPC2) channel. To investigate the physiological role played by the increase in intracellular calcium concentration in the apical region of these neurons, we produced localized, rapid, and reproducible increases in calcium concentration with flash photolysis of caged calcium and measured calcium-activated currents with the whole cell voltage-clamp technique. On average, a large inward calcium-activated current of −261 pA was measured at −50 mV, rising with a time constant of 13 ms. Ion substitution experiments showed that this current is anion selective. Moreover, the chloride channel blockers niflumic acid and 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid partially inhibited the calcium-activated current. These results directly demonstrate that a large chloride current can be activated by calcium in the apical region of mouse vomeronasal sensory neurons. Furthermore, we showed by immunohistochemistry that the calcium-activated chloride channels TMEM16A/anoctamin1 and TMEM16B/anoctamin2 are present in the apical layer of the vomeronasal epithelium, where they largely colocalize with the TRPC2 transduction channel. Immunocytochemistry on isolated vomeronasal sensory neurons showed that TMEM16A and TMEM16B coexpress in the neuronal microvilli. Therefore, we conclude that microvilli of mouse vomeronasal sensory neurons have a high density of calcium-activated chloride channels that may play an important role in vomeronasal transduction. PMID:22732308

  18. Calcium-activated chloride channels in the apical region of mouse vomeronasal sensory neurons.

    PubMed

    Dibattista, Michele; Amjad, Asma; Maurya, Devendra Kumar; Sagheddu, Claudia; Montani, Giorgia; Tirindelli, Roberto; Menini, Anna

    2012-07-01

    The rodent vomeronasal organ plays a crucial role in several social behaviors. Detection of pheromones or other emitted signaling molecules occurs in the dendritic microvilli of vomeronasal sensory neurons, where the binding of molecules to vomeronasal receptors leads to the influx of sodium and calcium ions mainly through the transient receptor potential canonical 2 (TRPC2) channel. To investigate the physiological role played by the increase in intracellular calcium concentration in the apical region of these neurons, we produced localized, rapid, and reproducible increases in calcium concentration with flash photolysis of caged calcium and measured calcium-activated currents with the whole cell voltage-clamp technique. On average, a large inward calcium-activated current of -261 pA was measured at -50 mV, rising with a time constant of 13 ms. Ion substitution experiments showed that this current is anion selective. Moreover, the chloride channel blockers niflumic acid and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid partially inhibited the calcium-activated current. These results directly demonstrate that a large chloride current can be activated by calcium in the apical region of mouse vomeronasal sensory neurons. Furthermore, we showed by immunohistochemistry that the calcium-activated chloride channels TMEM16A/anoctamin1 and TMEM16B/anoctamin2 are present in the apical layer of the vomeronasal epithelium, where they largely colocalize with the TRPC2 transduction channel. Immunocytochemistry on isolated vomeronasal sensory neurons showed that TMEM16A and TMEM16B coexpress in the neuronal microvilli. Therefore, we conclude that microvilli of mouse vomeronasal sensory neurons have a high density of calcium-activated chloride channels that may play an important role in vomeronasal transduction.

  19. Impact of calcium-activated potassium channels on NMDA spikes in cortical layer 5 pyramidal neurons

    PubMed Central

    Bock, Tobias

    2016-01-01

    Active electrical events play an important role in shaping signal processing in dendrites. As these events are usually associated with an increase in intracellular calcium, they are likely to be under the control of calcium-activated potassium channels. Here, we investigate the impact of calcium-activated potassium channels on N-methyl-d-aspartate (NMDA) receptor-dependent spikes, or NMDA spikes, evoked by glutamate iontophoresis onto basal dendrites of cortical layer 5 pyramidal neurons. We found that small-conductance calcium-activated potassium channels (SK channels) act to reduce NMDA spike amplitude but at the same time, also decrease the iontophoretic current required for their generation. This SK-mediated decrease in NMDA spike threshold was dependent on R-type voltage-gated calcium channels and indicates a counterintuitive, excitatory effect of SK channels on NMDA spike generation, whereas the capacity of SK channels to suppress NMDA spike amplitude is in line with the expected inhibitory action of potassium channels on dendritic excitability. Large-conductance calcium-activated potassium channels had no significant impact on NMDA spikes, indicating that these channels are either absent from basal dendrites or not activated by NMDA spikes. These experiments reveal complex and opposing interactions among NMDA receptors, SK channels, and voltage-gated calcium channels in basal dendrites of cortical layer 5 pyramidal neurons during NMDA spike generation, which are likely to play an important role in regulating the way these neurons integrate the thousands of synaptic inputs they receive. PMID:26936985

  20. The influence of discharge power and heat treatment on calcium phosphate coatings prepared by RF magnetron sputtering deposition.

    PubMed

    Yonggang, Yan; Wolke, J G C; Yubao, Li; Jansen, J A

    2007-06-01

    Ca-P coatings with different Ca/P ratio and composition were successfully prepared by RF magnetron sputtering deposition. The Ca/P ratio, phase composition, structure and morphological properties were characterized by XRD, FTIR, EDS and SEM analyses. All the as-sputtered coatings were amorphous and after IR-irradiation the coatings altered into a crystalline phase. The obtained coatings had a Ca/P ratio that varied from 0.55 to 2.10 and different phase compositions or mixtures of apatite, beta-pyrophosphate and beta-tricalciumphosphate structures were formed. Evidently, the phase compositions of the sputtered coatings are determined not only by the discharge power ratio of the hydroxylapatite and calcium pyrophosphate targets but also by the annealing temperature.

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

    SciTech Connect

    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.

  2. Carbon nanotubes play an important role in the spatial arrangement of calcium deposits in hydrogels for bone regeneration.

    PubMed

    Cancian, Giulia; Tozzi, Gianluca; Hussain, Amirul Ashraf Bin; De Mori, Arianna; Roldo, Marta

    2016-08-01

    Age related bone diseases such as osteoporosis are considered among the main causes of reduced bone mechanical stability and bone fractures. In order to restore both biological and mechanical function of diseased/fractured bones, novel bioactive scaffolds that mimic the bone structure are constantly under development in tissue engineering applications. Among the possible candidates, chitosan-based thermosensitive hydrogel scaffolds represent ideal systems due to their biocompatibility, biodegradability, enhanced antibacterial properties, promotion of osteoblast formation and ease of injection, which makes them suitable for less invasive surgical procedures. As a main drawback, these chitosan systems present poor mechanical performance that could not support load-bearing applications. In order to produce more mechanically-competent biomaterials, the combined addition of hydroxyapatite and carbon nanotubes (CNTs) is proposed in this study. Specifically, the aim of this work is to develop thermosensitive chitosan hydrogels containing stabilised single-walled and multi-walled CNTs, where their effect on the mechanical/physiochemical properties, calcium deposition patterns and ability to provide a platform for the controlled release of protein drugs was investigated. It was found that the addition of CNTs had a significant effect on the sol-gel transition time and significantly increased the resistance to compression for the hydrogels. Moreover, in vitro calcification studies revealed that CNTs played a major role in the spatial arrangements of newly formed calcium deposits in the composite materials studied, suggesting that they may have a role in the way the repair of fragile and/or fractured bones occurs in vivo.

  3. Controlled release based on the dissolution of a calcium carbonate layer deposited on hydrogels.

    PubMed

    Ogomi, Daisuke; Serizawa, Takeshi; Akashi, Mitsuru

    2005-03-21

    It is possible that inorganic materials conjugated to suitable organic materials may induce unique mechanical, optical and other functional properties. Therefore, artificial conjugation of organic and inorganic components is attractive for preparing novel functional materials. Recently, we developed an alternate soaking process for calcium salt formation on/in polymer materials. In this study, a poly(vinyl alcohol) (PVA) hydrogel-calcium carbonate (CaCO(3)) composite was prepared by the aforementioned process as a controlled release support. Brilliant blue FCF (Mw = 794), FITC labeled BSA (Mw = 6.6 x 10(4)), FITC labeled dextran-10 k (Mw = 9.5 x 10(3)) and FITC labeled dextran-40 k (Mw = 4.3 x 10(4)) were loaded into the composite as model drugs. CaCO(3) dissolution and model drug release rates increased with a decrease in buffer pH. In addition, model drug release rates increased with a decrease in model drug molecular weight. These results show that CaCO(3) layers on hydrogels behave as capping layers for model drug release; the release rate of model drugs can be controlled by the dissolution rate of CaCO(3) and the molecular weight of the drug.

  4. Requirement of calcium-activated chloride channels in the activation of mouse vomeronasal neurons

    PubMed Central

    Kim, SangSeong; Ma, Limei; Yu, C. Ron

    2011-01-01

    In terrestrial vertebrates, the vomeronasal organ (VNO) detects and transduces pheromone signals. VNO activation is thought to be mediated by the transient receptor potential C2 (TRPC2) channel. The aberrant behavioural phenotypes observed in TRPC2−/− mice are generally attributed to the lost VNO function. Recently, calcium-activated chloride channels have been shown to contribute to VNO activation. Here we show that CACCs can be activated in VNO slice preparations from the TRPC2−/− mice and this activation is blocked by pharmacological agents that inhibit intracellular Ca2+ release. Urine-evoked Cl− current is sufficient to drive spiking changes in VNO neurons from both wild-type (WT) and TRPC2−/− mice. Moreover, blocking Cl− conductance essentially abolishes VNO activation in WT neurons. These results suggest a TRPC2-independent signalling pathway in the VNO and the requirement of calcium-activated chloride channels currents to mediate pheromone activation. Our data further suggest that TRPC2−/− mice retain partial VNO function. PMID:21694713

  5. The Influence of Electrolytic Concentration on the Electrochemical Deposition of Calcium Phosphate Coating on a Direct Laser Metal Forming Surface

    PubMed Central

    Yang, Yuhui; Luo, Wenjing

    2017-01-01

    A calcium phosphate (CaP) coating on titanium surface enhances its biocompatibility, thus facilitating osteoconduction and osteoinduction with the inorganic phase of the human bone. Electrochemical deposition has been suggested as an effective means of fabricating CaP coatings on porous surface. The purpose of this study was to develop CaP coatings on a direct laser metal forming implant using electrochemical deposition and to investigate the effect of electrolytic concentration on the coating's morphology and structure by X-ray diffraction, scanning electron microscopy, water contact angle analysis, and Fourier transform infrared spectroscopy. In group 10−2, coatings were rich in dicalcium phosphate, characterized to be thick, layered, and disordered plates. In contrast, in groups 10−3 and 10−4, the relatively thin and well-ordered coatings predominantly consisted of granular hydroxyapatite. Further, the hydrophilicity and cell affinity were improved as electrolytic concentration increased. In particular, the cells cultured in group 10−3 appeared to have spindle morphology with thick pseudopodia on CaP coatings; these spindles and pseudopodia strongly adhered to the rough and porous surface. By analyzing and evaluating the surface properties, we provided further knowledge on the electrolytic concentration effect, which will be critical for improving CaP coated Ti implants in the future. PMID:28250771

  6. Soil calcium status and the response of stream chemistry to changing acidic deposition rates

    USGS Publications Warehouse

    Lawrence, G.B.; David, M.B.; Lovett, Gary M.; Murdoch, Peter S.; Burns, Douglas A.; Stoddard, J.L.; Baldigo, Barry P.; Porter, J.H.; Thompson, A.W.

    1999-01-01

    Despite a decreasing trend in acidic deposition rates over the past two to three decades, acidified surface waters in the northeastern United States have shown minimal changes. Depletion of soil Ca pools has been suggested as a cause, although changes in soil Ca pools have not been directly related to long-term records of stream chemistry. To investigate this problem, a comprehensive watershed study was conducted in the Neversink River Basin, in the Catskill Mountains of New York, during 1991-1996. Spatial variations of atmospheric deposition, soil chemistry, and stream chemistry were evaluated over an elevation range of 817-1234 m to determine whether these factors exhibited elevational patterns. An increase in atmospheric deposition of SO4 with increasing elevation corresponded with upslope decreases of exchangeable soil base concentrations and acid-neutralizing capacity of stream water. Exchangeable base concentrations in homogeneous soil incubated within the soil profile for one year also decreased with increasing elevation. An elevational gradient in precipitation was not observed, and effects of a temperature gradient on soil properties were not detected. Laboratory leaching experiments with soils from this watershed showed that (1) concentrations of Ca in leachate increased as the concentrations of acid anions in added solution increased, and (2) the slope of this relationship was positively correlated with base saturation. Field and laboratory soil analyses are consistent with the interpretation that decreasing trends in acid-neutralizing capacity in stream water in the Neversink Basin, dating back to 1984, are the result of decreases in soil base saturation caused by acidic deposition.

  7. Electromigration and Deposition of Micro-Scale Calcium Carbonate Structures with Controlled Morphology and Polymorphism

    DTIC Science & Technology

    2013-04-01

    this technique include synthesis of 3-D structures of mineral phases such as CaCO3 (e.g., biometic materials) or hydroxyapatite (e.g., artificial...infilling materials such as hydroxyapatite . ERDC/GSL TR-13-15 16 References Standard (ASTM). 2006. Standard test method for density, absorption, and...Journal of the European Ceramic Society 28 (7):1353-1367. Ma, J., C. Wang, and K. Peng. 2003. Electrophoretic deposition of porous hydroxyapatite

  8. Effects of Post-Deposition Annealing on the Properties of Calcium Manganese Oxide Thin Films

    NASA Astrophysics Data System (ADS)

    Ferrone, Natalie; Chaudhry, Adeel; Hart, Cacie; Lawson, Bridget; Houston, David; Neubauer, Samuel; Johnson, Anthony; Schaefer, David; Kolagani, Rajeswari

    We will present our results on the effects of post-deposition annealing on the structural and electrical properties of CaMnO3-d thin films grown by Pulsed Laser deposition. The thin films are epitaxially grown on (100) LaAlO3 which has larger in-plane lattice parameters than that of bulk CaMnO3, which leads to bi-axial tensile strain in the thin films. Results from our laboratory show that bi-axial tensile strain leads to low resistivity in thinner films, the resistivity increasing with increasing thickness. These results are suggestive of a coupling between strain and oxygen stoichiometry in the thin films. We have investigated the effects of post-deposition annealing in various gas ambients towards the goal of understanding the effects of relaxation and oxygen stoichiometric changes. We will present a comparison of the structural and electrical properties of as-grown and post-annealed films over a range of thicknesses. Support from Towson University Office of Undergraduate Research, Fisher Endowment Grant & Undergraduate Research Grant from the Fisher College of Science & Mathematics, Seed Funding Grant from the School of Emerging technologies, & NSF Grant ECCS 112856.

  9. Detection of calcium activity in human monocytes by the fura-2 fluorescence method: in vitro differentiation sensitizes cells to dihydropyridine calcium channel modulators

    NASA Astrophysics Data System (ADS)

    Oraevsky, Alexander A.; Cabello, Olga A.; Shan, Qin; Tittel, Frank K.; Henry, Philip D.

    1994-07-01

    Dihydropyridine (DHP) calcium channel blockers have been shown to suppress atherogenesis in various species and controlled angiographic trials suggest that these drugs may retard the progression of occlusive coronary disease in humans. Because mononuclear leukocytes play a key role in the formation of early and advanced atheromatous lesions, we determined effects of DHP calcium channel modulators on calcium uptake by cells of the monocytic lineage. Human peripheral blood monocytes were evaluated before and after undergoing in vitro differentiation induced by two days of culture with fetal calf serum and FMLP. Changes in intracellular calcium activity were estimated with fura-2, a fluorescent calcium indicator. Freshly isolated (unactivated) monocytes were insensitive to DHP drugs both in the presence and absence of high potassium membrane depolarization. In contrast, nisoldipine, a DHP calcium channel blocker, and BAY K 8644, a DHP calcium channel activator, decreased and increased calcium uptake by KC1-depolarized differentiated monocytes. Results suggest that differentiation of monocytes to macrophages may involve a change in the expression and/or regulation of DHP- sensitive calcium channels.

  10. Calcium mobilization and phosphoinositide turnover in fluoride-activated human neutrophils

    SciTech Connect

    Strnad, C.F.; Wong, K.

    1986-05-01

    Fluoride ion, at concentrations above 10 mM, has been found to activate a superoxide production response in human neutrophils which is strongly dependent on the presence of extracellular calcium. In an attempt to further explore the calcium requirement of fluoride-induced neutrophil activation, intracellular calcium concentrations were monitored through use of the fluorescent calcium probe, Quin 2. Fluoride ion, at concentrations between 10 and 20 mM, was found to elicit a rise in intracellular calcium levels which was characterized by a lag period of 4 to 10 min and a prolonged duration of action (greater than 20 min). In contrast, the chemotactic peptide, formylmethionyl-leucyl-phenylalanine (FMLP), induced a rise in intracellular calcium concentration which peaked within 1 min. Preincubation of cells with 1 ..mu..g/ml pertussis toxin resulted in inhibition of the FMLP-induced response, but not that elicited by fluoride. Furthermore, anion exchange chromatography indicated that inositol phosphate accumulation occurred in fluoride-treated cells in association with calcium mobilization. Recent evidence suggests that the FMLP receptor is coupled to phospholipase C and phosphoinositide turnover through a guanine nucleotide binding protein susceptible to inhibition by pertussis toxin. Present results suggest that fluoride ion may serve to activate this protein in a manner resistant to inhibition by pertussis toxin.

  11. Activation of protein kinase C inhibits calcium-activated potassium channels in rat pituitary tumour cells.

    PubMed Central

    Shipston, M J; Armstrong, D L

    1996-01-01

    1. The regulation of large-conductance, calcium- and voltage-dependent potassium (BK) channels by protein kinase C (PKC) was investigated in clonal rat anterior pituitary cells (GH4C1), which were voltage clamped at -40 mV in a physiological potassium gradient through amphotericin-perforated patches. 2. Maximal activation of PKC by 100 nM phorbol 12, 13-dibutyrate (PdBu) almost completely inhibited the voltage-activated outward current through BK channels. In contrast PdBu had no significant effect on the residual outward current after block of BK channels with 2 mM TEA or 30 nM charybdotoxin. In single-channel recordings from cell-attached patches, PdBu reduced the open probability of BK channels more than eightfold with no significant effect on mean open lifetime or unitary conductance. 3. The effects of PdBu on BK channels were not mimicked by the 4 alpha-isomer, which does not activate PKC, and were blocked almost completely by 25 microM chelerythrine, a specific, noncompetitive PKC inhibitor. 4. PdBu had no significant effect on the amplitude of the pharmacologically isolated, high voltage-activated calcium current. 5. Inhibition of BK channel activity by PKC provides the first molecular mechanism linking hormonal activation of phospholipase C to sustained excitability in pituitary cells. PMID:8799890

  12. Change perspective to increase diagnostic accuracy of ultrasonography in calcium pyrophosphate dehydrate deposition disease! A new approach: the axial scan of the meniscus.

    PubMed

    Filippou, G; Picerno, V; Adinolfi, A; Di Sabatino, V; Bertoldi, I; Galeazzi, M; Frediani, B

    2015-03-31

    Ultrasonography (US) is a relevant tool in the study of calcium pyrophosphate dihydrate (CPP) deposition disease. However, differential diagnosis of hyperechoic deposits within the fibrocartilage can be difficult; moreover, US study is limited by the need of an adequate acoustic window. We describe a US scanning technique that offers a new viewpoint in the study of knee meniscal structure: a longitudinal scan performed according to the long axis of meniscus. This technique proves to be particularly useful for the identification of CPP deposition, but could also improve the US diagnostic utility and accuracy in other meniscal pathologies.

  13. The prevalence of chondrocalcinosis of the symphysis pubis on CT scan and correlation with calcium pyrophosphate dihydrate crystal deposition disease.

    PubMed

    Patel, Trusha; Ryan, Lawrence; Dubois, Melissa; Carrera, Guillermo; Baynes, Keith; Mannem, Rajeev; Mulkerin, Jennifer; Visotcky, Alexis

    2016-03-01

    Calcium pyrophosphate dihydrate (CPP) crystal deposition in the articular cartilage can often be seen radiographically as chondrocalcinosis (CC). CPP crystals preferentially deposit in fibrocartilages such as the knee menisci and symphysis pubis (SP). We sought to determine the prevalence of CC in the SP on computed tomography (CT) of the abdomen and pelvis. This retrospective study involved readings on 1070 consecutive CTs of the abdomen and pelvis performed over 3 months in patients over 65 years of age. Medical records of 226 patients found to have CC were reviewed to determine age, gender, documentation of CPPD on problem lists or in medical histories, and whether radiology readings of the CTs mentioned CC. SP CC was identified in 21.1 % (226/1070) of consecutive CT scans with the mean age of CT+ patients being 78.6. Of the 226 patients with SP CC, the observation of CC was documented in only 5.3 % (12/226) of the radiology reports. Of the 12 instances in which the radiology reports mentioned CC, this observation was never (0/12) transmitted to the medical history or problem list. The prevalence of SP CC in patients older than 65 was 21.1 %. Since the majority of CTs of the abdomen and pelvis are not ordered for evaluation of musculoskeletal conditions, this is likely a true prevalence without selection bias. When CC of the SP was present on images, radiologists routinely overlooked or chose not to report CC. Even in the rare instances when it was reported, that information was not added to the medical history or problem list. There are several clinical situations (e.g., acute monoarthritis or atypical osteoarthritis) in which recognizing that a patient has CPP deposition would be useful. Taking the time to review images may yield clinically important findings that are not mentioned anywhere on the patient chart.

  14. Role of calcium and calcium-activated proteases in CYP2E1-dependent toxicity in HEPG2 cells.

    PubMed

    Caro, Andres A; Cederbaum, Arthur I

    2002-01-04

    The objective of this work was to investigate whether CYP2E1- and oxidative stress-dependent toxicity in HepG2 cells is mediated by an increase of cytosolic Ca2+ and activation of Ca2+-modulated processes. HepG2 cells expressing CYP2E1 (E47 cells) or control cells not expressing CYP2E1 (C34 cells) were preloaded with arachidonic acid (AA, up to 10 microm) and, after washing, incubated with iron-nitrilotriacetic acid (up to 100 microm) for variable periods (up to 12 h). Toxicity was greater in E47 cells than in C34 cells at all times and combinations of iron/AA tested. Cytosolic calcium increased with incubation time in both cell lines, but the increase was higher in E47 cells than in C34 cells. The rise in calcium was an early event and preceded the developing toxicity. Toxicity in E47 cells and the increase in Ca2+ were inhibited by omission of Ca2+ from the extracellular medium, and toxicity was restored by reincorporation of Ca2+. An inhibitor of Ca2+ release from intracellular stores did not prevent the toxicity or the increase in Ca2+, reflecting a role for the influx of extracellular Ca2+ in the toxicity. Reactive oxygen production was similar in media with or without calcium, indicating that calcium was not modulating CYP2E1-dependent oxidative stress. Toxicity, lipid peroxidation, and the increase of Ca2+ in E47 cells exposed to iron-AA were inhibited by alpha-tocopherol. E47 cells (but not C34 cells) exposed to iron-AA showed increased calpain activity in situ (40-fold). The toxicity in E47 cells mirrored calpain activation and was inhibited by calpeptin, suggesting that calpain activation plays a causal role in toxicity. These results suggest that CYP2E1-dependent toxicity in this model depends on the activation of lipid peroxidation, followed by an increased influx of extracellular Ca2+ and activation of Ca2+-dependent proteases.

  15. Antagonists of the TMEM16A Calcium-Activated Chloride Channel Modulate Airway Smooth Muscle Tone and Intracellular Calcium

    PubMed Central

    Danielsson, Jennifer; Perez-Zoghbi, Jose; Bernstein, Kyra; Barajas, Matthew B.; Zhang, Yi; Kumar, Satish; Sharma, Pawan K.; Gallos, George; Emala, Charles W.

    2015-01-01

    Background Perioperative bronchospasm refractory to β-agonists continues to challenge anesthesiologists and intensivists. The TMEM16A calcium-activated chloride channel modulates airway smooth muscle (ASM) contraction. We hypothesized that TMEM16A antagonists would relax ASM contraction by modulating membrane potential and calcium flux. Methods Human ASM, guinea pig tracheal rings or mouse peripheral airways were contracted with acetylcholine (Ach) or leukotriene D4 (LTD4) and then treated with the TMEM16A antagonists: benzbromarone, T16Ainh-A01, MONNA or B25. In separate studies, guinea pig tracheal rings were contracted with Ach and then exposed to increasing concentrations of isoproterenol (0.01nM-10μM) ± benzbromarone. Plasma membrane potential and intracellular calcium concentrations were measured in human ASM cells. Results Benzbromarone was the most potent TMEM16A antagonist tested for relaxing an Ach-induced contraction in guinea pig tracheal rings (n=6). Further studies were done to investigate benzbromarone’s clinical utility. In human ASM, benzbromarone relaxed either an acetylcholine- or LTD4-induced contraction (n=8). Benzbromarone was also effective in relaxing peripheral airways (n=9) and potentiating relaxation by β-agonists (n=5–10). In cellular mechanistic studies, benzbromarone hyperpolarized human ASM cells (n=9–12) and attenuated intracellular calcium flux from both the plasma membrane and sarcoplasmic reticulum (n=6–12). Conclusions TMEM16A antagonists work synergistically with β-agonists and through a novel pathway of interrupting ion flux both at the plasma membrane and sarcoplasmic reticulum to acutely relax human airway smooth muscle. PMID:26181339

  16. Levetiracetam Inhibits Both Ryanodine and IP3 Receptor Activated Calcium Induced Calcium Release in Hippocampal Neurons in Culture

    PubMed Central

    Nagarkatti, Nisha; Deshpande, Laxmikant S.; DeLorenzo, Robert J.

    2010-01-01

    Epilepsy affects approximately 1% of the population worldwide, and there is a pressing need to develop new anti-epileptic drugs (AEDs) and understand their mechanisms of action. Levetiracetam (LEV) is a novel AED and despite its increasingly widespread clinical use, its mechanism of action is as yet undetermined. Intracellular calcium ([Ca2+]i) regulation by both inositol 1,4,5-triphosphate receptors (IP3R) and ryanodine receptors (RyR) has been implicated in epileptogenesis and the maintenance of epilepsy. To this end, we investigated the effect of LEV on RyR and IP3R activated calcium-induced calcium release (CICR) in hippocampal neuronal cultures. RyR-mediated CICR was stimulated using the well-characterized RyR activator, caffeine. Caffeine (10mM) caused a significant increase in [Ca2+]i in hippocampal neurons. Treatment with LEV (33μM) prior to stimulation of RyR-mediated CICR by caffeine led to a 61% decrease in the caffeine induced peak height of [Ca2+]i when compared to the control. Bradykinin stimulates IP3R-activated CICR—to test the effect of LEV on IP3R-mediated CICR, bradykinin (1μM) was used to stimulate cells pre-treated with LEV (100μM). The data showed that LEV caused a 74% decrease in IP3R-mediated CICR compared to the control. In previous studies we have shown that altered Ca2+ homeostatic mechanisms play a role in seizure activity and the development of spontaneous recurrent epileptiform discharges (SREDs). Elevations in [Ca2+]i mediated by CICR systems have been associated with neurotoxicity, changes in neuronal plasticity, and the development of AE. Thus, the ability of LEV to modulate the two major CICR systems demonstrates an important molecular effect of this agent on a major second messenger system in neurons. PMID:18406528

  17. Inhibition of calcium carbonate deposition on stainless steel using olive leaf extract as a green inhibitor.

    PubMed

    Aidoud, Roumaissa; Kahoul, Abdelkrim; Naamoune, Farid

    2017-01-01

    The antiscale properties of the aqueous extract of olive (Olea europaea L.) leaves as a natural scale inhibitor for stainless steel surface in Hammam raw water were investigated using chronoamperometry (CA) and electrochemical impedance spectroscopy techniques in conjunction with a microscopic examination. The X-ray diffraction analysis reveals that the scale deposited over the pipe walls consists of pure CaCO3 calcite. The CA, in accordance with electrochemical impedance spectra and scanning electron microscopy, shows that the inhibition efficiency increases with increasing extract concentration. This efficiency is considerably reduced as the temperature is increased.

  18. Synthesis, Evaluation of Pharmacological Activity, and Molecular Docking of 1,4-Dihydropyridines as Calcium Antagonists.

    PubMed

    Shaldam, Moataz Ahmed; El-Hamamsy, Mervat Hamed; Saleh, Dalia Osama; El-Moselhy, Tarek Fathy

    2016-01-01

    1,4-Dihydropyridine (DHP) is an important class of calcium antagonist. It inhibits the influx of extracellular Ca(2+) through L-type voltage-dependent calcium channels. Two series of nifedipine analogues were synthesized and evaluated as calcium antagonists. The ortho-nitrophenyl ring of nifedipine was replaced with an ortho- or a meta-chlorophenyl substituent. The IC50 values revealed that some of the compounds are similar to or more active than nifedipine. Substitution with groups of suitable bulkiness, such as ethyl ester, at the 3- and 5-positions of the DHP ring gave 3h, which is approximately three-fold more active than nifedipine as a calcium antagonist. A docking study with the DHP receptor model was performed to interpret the differences in calcium antagonist activities. The molecular docking study demonstrated that the lipophilicity of the substituted phenyl group at the 4-position of the DHP ring is an important factor that could increase the activity of the calcium antagonist taking the steric factor into consideration. Bulky groups interfere with ring-to-ring hydrophobic interaction with Tyr(1460) and limit the efficiency of increasing the length of the hydrocarbon chain of esters at the 3- and 5-positions of the DHP ring as an approach to increase activity. The presence of a chelating substituent on the phenyl ring at the 4-position of the DHP ring may ensure strong binding to the receptor and hence stabilization of the closed-channel conformation.

  19. Hydrogen peroxide homeostasis: activation of plant catalase by calcium/calmodulin

    NASA Technical Reports Server (NTRS)

    Yang, T.; Poovaiah, B. W.

    2002-01-01

    Environmental stimuli such as UV, pathogen attack, and gravity can induce rapid changes in hydrogen peroxide (H(2)O(2)) levels, leading to a variety of physiological responses in plants. Catalase, which is involved in the degradation of H(2)O(2) into water and oxygen, is the major H(2)O(2)-scavenging enzyme in all aerobic organisms. A close interaction exists between intracellular H(2)O(2) and cytosolic calcium in response to biotic and abiotic stresses. Studies indicate that an increase in cytosolic calcium boosts the generation of H(2)O(2). Here we report that calmodulin (CaM), a ubiquitous calcium-binding protein, binds to and activates some plant catalases in the presence of calcium, but calcium/CaM does not have any effect on bacterial, fungal, bovine, or human catalase. These results document that calcium/CaM can down-regulate H(2)O(2) levels in plants by stimulating the catalytic activity of plant catalase. Furthermore, these results provide evidence indicating that calcium has dual functions in regulating H(2)O(2) homeostasis, which in turn influences redox signaling in response to environmental signals in plants.

  20. Localization of calcium stimulated adenosine triphosphatase activity in blood vessels of the skeleton

    NASA Technical Reports Server (NTRS)

    Doty, S. B.

    1985-01-01

    Alkaline phosphatase is an enzyme found in bone forming cells which decreases in certain bones as a result of hypogravity or non-weight bearing. This enzyme can also hydrolyze adenosine triphosphate. Therefore, an effort was made to localize calcium-stimulated ATPase by cytochemistry to determine whether altered bone cell activity might be related to changing calcium levels which occur during hypogravity. The results indicate that Ca(++)-ATPase is largely found along the endothelium and basal lamina of blood vessels, and not found in bone forming cells. This suggests that calcium regulation in the vicinity of bone formation may be modulated by the vasculature of the area.

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

    PubMed

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

    2013-04-15

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

  2. Structure-activity relationship study of 1,4-dihydropyridine derivatives blocking N-type calcium channels.

    PubMed

    Yamamoto, Takashi; Niwa, Seiji; Ohno, Seiji; Onishi, Tomoyuki; Matsueda, Hiroyuki; Koganei, Hajime; Uneyama, Hisayuki; Fujita, Shin-ichi; Takeda, Tomoko; Kito, Morikazu; Ono, Yukitsugu; Saitou, Yuki; Takahara, Akira; Iwata, Seinosuke; Shoji, Masataka

    2006-02-15

    Cilnidipine is a 1,4-dihydropyridine derived L/N-type calcium channel dual blocker possessing neuroprotective and analgesic effects which are related to its N-type calcium channel inhibitory activity. In order to find specific N-type calcium channel blockers with the least effects on cardiovascular system, we performed structure-activity relationship study on APJ2708, which is a derivative of cilnidipine, and found a promising N-type calcium channel blocker 21b possessing analgesic effect in vivo with a 1600-fold lower activity against L-type calcium channels than that of cilnidipine.

  3. Calcium-activated potassium conductance noise in snail neurons.

    PubMed

    Westerfield, M; Lux, H D

    1982-11-01

    Current fluctuations were measured in small, 3-6 micrometers-diameter patches of soma membrane in bursting neurons of the snail, Helix pomatia. The fluctuations dramatically increased in magnitude with depolarization of the membrane potential under voltage clamp conditions. Two components of conductance noise were identified in the power spectra calculated from the membrane currents. One component had a corner frequency which increased with depolarization. This component was blocked by intracellular injection of TEA and was relatively insensitive to extracellular calcium levels (as long as the total number of effective divalent cations remained constant). It was identified as fluctuations of the voltage-dependent component of delayed outward current. The second component of conductance noise had a corner frequency which decreased with depolarization. It was relatively unaffected by TEA injection and was reversibly blocked by substitution of extracellular calcium with magnesium, cobalt, or nickel. This second component of noise was identified as fluctuations of the calcium-dependent potassium current. The results suggest that the two components of delayed outward current are conducted through physically distinct channels.

  4. Role of calcium signaling in the activation of mitochondrial nitric oxide synthase and citric acid cycle.

    PubMed

    Traaseth, Nathaniel; Elfering, Sarah; Solien, Joseph; Haynes, Virginia; Giulivi, Cecilia

    2004-07-23

    An apparent discrepancy arises about the role of calcium on the rates of oxygen consumption by mitochondria: mitochondrial calcium increases the rate of oxygen consumption because of the activation of calcium-activated dehydrogenases, and by activating mitochondrial nitric oxide synthase (mtNOS), decreases the rates of oxygen consumption because nitric oxide is a competitive inhibitor of cytochrome oxidase. To this end, the rates of oxygen consumption and nitric oxide production were followed in isolated rat liver mitochondria in the presence of either L-Arg (to sustain a mtNOS activity) or N(G)-monomethyl-L-Arg (NMMA, a competitive inhibitor of mtNOS) under State 3 conditions. In the presence of NMMA, the rates of State 3 oxygen consumption exhibited a K(0.5) of 0.16 microM intramitochondrial free calcium, agreeing with those required for the activation of the Krebs cycle. By plotting the difference between the rates of oxygen consumption in State 3 with L-Arg and with NMMA at various calcium concentrations, a K(0.5) of 1.2 microM intramitochondrial free calcium was obtained, similar to the K(0.5) (0.9 microM) of the dependence of the rate of nitric oxide production on calcium concentrations. The activation of dehydrogenases, followed by the activation of mtNOS, would lead to the modulation of the Krebs cycle activity by the modulation of nitric oxide on the respiratory rates. This would ensue in changes in the NADH/NAD and ATP/ADP ratios, which would influence the rate of the cycle and the oxygen diffusion.

  5. PKCα mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells

    PubMed Central

    Adapala, Ravi K.; Talasila, Phani K.; Bratz, Ian N.; Zhang, David X.; Suzuki, Makoto; Meszaros, J. Gary

    2011-01-01

    Transient receptor potential vanilloid channel 4 (TRPV4) is a polymodally activated nonselective cationic channel implicated in the regulation of vasodilation and hypertension. We and others have recently shown that cyclic stretch and shear stress activate TRPV4-mediated calcium influx in endothelial cells (EC). In addition to the mechanical forces, acetylcholine (ACh) was shown to activate TRPV4-mediated calcium influx in endothelial cells, which is important for nitric oxide-dependent vasodilation. However, the molecular mechanism through which ACh activates TRPV4 is not known. Here, we show that ACh-induced calcium influx and endothelial nitric oxide synthase (eNOS) phosphorylation but not calcium release from intracellular stores is inhibited by a specific TRPV4 antagonist, AB-159908. Importantly, activation of store-operated calcium influx was not altered in the TRPV4 null EC, suggesting that TRPV4-dependent calcium influx is mediated through a receptor-operated pathway. Furthermore, we found that ACh treatment activated protein kinase C (PKC) α, and inhibition of PKCα activity by the specific inhibitor Go-6976, or expression of a kinase-dead mutant of PKCα but not PKCε or downregulation of PKCα expression by chronic 12-O-tetradecanoylphorbol-13-acetate treatment, completely abolished ACh-induced calcium influx. Finally, we found that ACh-induced vasodilation was inhibited by the PKCα inhibitor Go-6976 in small mesenteric arteries from wild-type mice, but not in TRPV4 null mice. Taken together, these findings demonstrate, for the first time, that a specific isoform of PKC, PKCα, mediates agonist-induced receptor-mediated TRPV4 activation in endothelial cells. PMID:21705673

  6. Basal and Activated Calcium Sensitization Mediated by RhoA/Rho Kinase Pathway in Rats with Genetic and Salt Hypertension

    PubMed Central

    Bencze, Michal; Vaněčková, Ivana; Kuneš, Jaroslav; Zicha, Josef

    2017-01-01

    Calcium sensitization mediated by RhoA/Rho kinase pathway can be evaluated either in the absence (basal calcium sensitization) or in the presence of endogenous vasoconstrictor systems (activated calcium sensitization). Our aim was to compare basal and activated calcium sensitization in three forms of experimental hypertension with increased sympathetic tone and enhanced calcium entry—spontaneously hypertensive rats (SHR), heterozygous Ren-2 transgenic rats (TGR), and salt hypertensive Dahl rats. Activated calcium sensitization was determined as blood pressure reduction induced by acute administration of Rho kinase inhibitor fasudil in conscious rats with intact sympathetic nervous system (SNS) and renin-angiotensin system (RAS). Basal calcium sensitization was studied as fasudil-dependent difference in blood pressure response to calcium channel opener BAY K8644 in rats subjected to RAS and SNS blockade. Calcium sensitization was also estimated from reduced development of isolated artery contraction by Rho kinase inhibitor Y-27632. Activated calcium sensitization was enhanced in all three hypertensive models (due to the hyperactivity of vasoconstrictor systems). In contrast, basal calcium sensitization was reduced in SHR and TGR relative to their controls, whereas it was augmented in salt-sensitive Dahl rats relative to their salt-resistant controls. Similar differences in calcium sensitization were seen in femoral arteries of SHR and Dahl rats. PMID:28197417

  7. Basal and Activated Calcium Sensitization Mediated by RhoA/Rho Kinase Pathway in Rats with Genetic and Salt Hypertension.

    PubMed

    Behuliak, Michal; Bencze, Michal; Vaněčková, Ivana; Kuneš, Jaroslav; Zicha, Josef

    2017-01-01

    Calcium sensitization mediated by RhoA/Rho kinase pathway can be evaluated either in the absence (basal calcium sensitization) or in the presence of endogenous vasoconstrictor systems (activated calcium sensitization). Our aim was to compare basal and activated calcium sensitization in three forms of experimental hypertension with increased sympathetic tone and enhanced calcium entry-spontaneously hypertensive rats (SHR), heterozygous Ren-2 transgenic rats (TGR), and salt hypertensive Dahl rats. Activated calcium sensitization was determined as blood pressure reduction induced by acute administration of Rho kinase inhibitor fasudil in conscious rats with intact sympathetic nervous system (SNS) and renin-angiotensin system (RAS). Basal calcium sensitization was studied as fasudil-dependent difference in blood pressure response to calcium channel opener BAY K8644 in rats subjected to RAS and SNS blockade. Calcium sensitization was also estimated from reduced development of isolated artery contraction by Rho kinase inhibitor Y-27632. Activated calcium sensitization was enhanced in all three hypertensive models (due to the hyperactivity of vasoconstrictor systems). In contrast, basal calcium sensitization was reduced in SHR and TGR relative to their controls, whereas it was augmented in salt-sensitive Dahl rats relative to their salt-resistant controls. Similar differences in calcium sensitization were seen in femoral arteries of SHR and Dahl rats.

  8. Calcium uptake and ATPase activity of sarcoplasmic reticulum vesicles isolated from control and selenium deficient lambs.

    PubMed

    Tripp, M J; Whanger, P D; Schmitz, J A

    1993-06-01

    The calcium uptake and ATPase activity were studied using fragmented sarcoplasmic reticulum (FSR) vesicles from normal and selenium (vitamin E)--deficient lambs. The latter group was suffering from white muscle disease (WMD). The calcium uptake of FSR vesicles from muscle of WMD lambs was reduced 10-fold as compared to those from normal lambs. An inverse relationship was found with the calcium uptake ability of the FSR vesicles and the severity of WMD. ATPase activity was nonsignificantly lower in vesicles from WMD lambs. The most active FSR vesicles from both normal and WMD lambs banded at 27% when purified on linear sucrose density gradients. The number of protein bands appearing in acrylamide gels of the purified vesicles appeared to be directly proportional to the severity of WMD. The 75Se cosedimented with the calcium uptake and ATPase activity when FSR vesicles from a lamb injected with 75Se-selenite were subjected to linear sucrose density gradient centrifugation, suggesting that selenium is incorporated into these vesicles. Injection of selenium into WMD lambs resulted in significantly greater calcium uptake activity in vesicles 18 and 38 days later as compared with untreated WMD lambs. Injection of selenium in WMD lambs resulted in a marked decrease in plasma CPK activity and a significant increase of glutathione peroxidase activity in the blood.

  9. Discovery, structure-activity relationship study, and oral analgesic efficacy of cyproheptadine derivatives possessing N-type calcium channel inhibitory activity.

    PubMed

    Yamamoto, Takashi; Niwa, Seiji; Iwayama, Satoshi; Koganei, Hajime; Fujita, Shin-ichi; Takeda, Tomoko; Kito, Morikazu; Ono, Yukitsugu; Saitou, Yuki; Takahara, Akira; Iwata, Seinosuke; Yamamoto, Hiroshi; Shoji, Masataka

    2006-08-01

    Antiallergic drug cyproheptadine (Cyp) is known to have inhibitory activities for L-type calcium channels in addition to histamine and serotonin receptors. Since we found that Cyp had an inhibitory activity against N-type calcium channel, Cyp was optimized to obtain more selective N-type calcium channel blocker with analgesic action. As a consequence of the optimization, we found 13 with potent N-type calcium channel inhibitory activity which had lower inhibitory activities against L-type calcium channel, histamine (H1), and serotonin (5-HT2A) receptors than those of Cyp. 13 showed an oral analgesic activity in rat formalin-induced pain model.

  10. Calcium-activated butyrylcholinesterase in human skin protects acetylcholinesterase against suicide inhibition by neurotoxic organophosphates

    SciTech Connect

    Schallreuter, Karin U.; University of Bradford ). E-mail: K.Schallreuter@bradford.ac.uk; Gibbons, Nicholas C.J.; Elwary, Souna M.; Parkin, Susan M.; Wood, John M.

    2007-04-20

    The human epidermis holds an autocrine acetylcholine production and degradation including functioning membrane integrated and cytosolic butyrylcholinesterase (BuchE). Here we show that BuchE activities increase 9-fold in the presence of calcium (0.5 x 10{sup -3}M) via a specific EF-hand calcium binding site, whereas acetylcholinesterase (AchE) is not affected. {sup 45}Calcium labelling and computer simulation confirmed the presence of one EF-hand binding site per subunit which is disrupted by H{sub 2}O{sub 2}-mediated oxidation. Moreover, we confirmed the faster hydrolysis by calcium-activated BuchE using the neurotoxic organophosphate O-ethyl-O-(4-nitrophenyl)-phenylphosphonothioate (EPN). Considering the large size of the human skin with 1.8 m{sup 2} surface area with its calcium gradient in the 10{sup -3}M range, our results implicate calcium-activated BuchE as a major protective mechanism against suicide inhibition of AchE by organophosphates in this non-neuronal tissue.

  11. Wet deposition of the seeding agent after weather modification activities.

    PubMed

    Curić, Mladjen; Janc, Dejan

    2013-09-01

    Weather modification activities are performed mostly by cloud seeding. Some operational projects have been conducted for more than a half century and cover planetary scales. These activities have led to large amounts of seeding agents being deposited on the ground in precipitation. The main intent of this paper is to identify the spatial pattern of silver iodide deposits after hail suppression. The spatial pattern of silver iodide deposits is determined using the weather modification project measurements from seeding agent reports, two weather radars and 316 launching sites during a 5-year period. The estimated spatial distribution of the deposits is not uniform, with the maximum silver iodide amount located in the southern part of the study area (up to 140 μg m(-2)). Our results are comparable with the measurements performed by chemical analyses during other cloud seeding experiments. The maximum location coincides well with that of the maximum seeded hailstorm precipitation frequency. A new method for identifying the spatial pattern of wet-deposited material has been established. The location with the maximum amount is found. This method would be important as a means of placing samplers and monitoring at the representative sites because those are where most weather modification projects would be performed in the future.

  12. Tooth development in Ambystoma mexicanum: phosphatase activities, calcium accumulation and cell proliferation in the tooth-forming tissues.

    PubMed

    Wistuba, Joachim; Ehmcke, Jens; Clemen, Günter

    2003-06-01

    Prerequisites of tooth formation, cell proliferation in the tooth-forming tissues, calcium accumulation and the enzymatic activities of alkaline (ALP) and acid phosphatases (ACP) were investigated by immunohistochemical and histochemical methods in various developmental stages of the Mexican Axolotl, Ambystoma mexicanum. During the growth of replacement teeth, the tooth-forming tissues continually recruit cells from the surrounding regions. The basal layer of the oral epithelium, the dental lamina and sometimes even the outer enamel epithelium provide cells for the differentiated inner enamel epithelium, in which the active ameloblasts are localized. The differentiating odontoblasts are derived from proliferating cells situated basally to the replacement teeth in the mesenchymal tissue. When differentiation has started and the cells have become functional, proliferative activity can no longer be observed. Calcium is accumulated close to the site of mineralization in the inner enamel epithelium and in the odontoblasts as it is in mammals, elasmobranchii and teleostei. The activities of ACP and ALP related to the mineralization of the replacement teeth are separated spatially and not sequentially as they are in mammals. However, the results indicate a similar function of these enzymatic components in relation to tooth formation and maturation of mineral deposition. Most of the substantial processes related to tooth formation reported from other vertebrates occur in a manner similar to that in Ambystoma mexicanum, but there also seem to be basic mechanisms present that are realised in a unique way in this urodele.

  13. Calcium micro-depositions in jugular truncular venous malformations revealed by Synchrotron-based XRF imaging.

    PubMed

    Pascolo, Lorella; Gianoncelli, Alessandra; Rizzardi, Clara; Tisato, Veronica; Salomé, Murielle; Calligaro, Carla; Salvi, Fabrizio; Paterson, David; Zamboni, Paolo

    2014-10-07

    It has been recently demonstrated that the internal jugular vein may exhibit abnormalities classified as truncular venous malformations (TVMs). The investigation of possible morphological and biochemical anomalies at jugular tissue level could help to better understand the link between brain venous drainage and neurodegenerative disorders, recently found associated with jugular TVMs. To this end we performed sequential X-ray Fluorescence (XRF) analyses on jugular tissue samples from two TVM patients and two control subjects, using complementary energies at three different synchrotrons. This investigation, coupled with conventional histological analyses, revealed anomalous micro-formations in the pathological tissues and allowed the determination of their elemental composition. Rapid XRF analyses on large tissue areas at 12.74 keV showed an increased Ca presence in the pathological samples, mainly localized in tunica adventitia microvessels. Investigations at lower energy demonstrated that the high Ca level corresponded to micro-calcifications, also containing P and Mg. We suggest that advanced synchrotron XRF micro-spectroscopy is an important analytical tool in revealing biochemical changes, which cannot be accessed by conventional investigations. Further research on a larger number of samples is needed to understand the pathogenic significance of Ca micro-depositions detected on the intramural vessels of vein walls affected by TVMs.

  14. Fast activation of dihydropyridine-sensitive calcium channels of skeletal muscle. Multiple pathways of channel gating

    PubMed Central

    1996-01-01

    Dihydropyridine (DHP) receptors of the transverse tubule membrane play two roles in excitation-contraction coupling in skeletal muscle: (a) they function as the voltage sensor which undergoes fast transition to control release of calcium from sarcoplasmic reticulum, and (b) they provide the conducting unit of a slowly activating L-type calcium channel. To understand this dual function of the DHP receptor, we studied the effect of depolarizing conditioning pulse on the activation kinetics of the skeletal muscle DHP-sensitive calcium channels reconstituted into lipid bilayer membranes. Activation of the incorporated calcium channel was imposed by depolarizing test pulses from a holding potential of -80 mV. The gating kinetics of the channel was studied with ensemble averages of repeated episodes. Based on a first latency analysis, two distinct classes of channel openings occurred after depolarization: most had delayed latencies, distributed with a mode of 70 ms (slow gating); a small number of openings had short first latencies, < 12 ms (fast gating). A depolarizing conditioning pulse to +20 mV placed 200 ms before the test pulse (-10 mV), led to a significant increase in the activation rate of the ensemble averaged-current; the time constant of activation went from tau m = 110 ms (reference) to tau m = 45 ms after conditioning. This enhanced activation by the conditioning pulse was due to the increase in frequency of fast open events, which was a steep function of the intermediate voltage and the interval between the conditioning pulse and the test pulse. Additional analysis demonstrated that fast gating is the property of the same individual channels that normally gate slowly and that the channels adopt this property after a sojourn in the open state. The rapid secondary activation seen after depolarizing prepulses is not compatible with a linear activation model for the calcium channel, but is highly consistent with a cyclical model. A six- state cyclical model is

  15. Activation of TRPV1-dependent calcium oscillation exacerbates seawater inhalation-induced acute lung injury

    PubMed Central

    LI, CONGCONG; BO, LIYAN; LIU, QINGQING; LIU, WEI; CHEN, XIANGJUN; XU, DUNQUAN; JIN, FAGUANG

    2016-01-01

    Calcium is an important second messenger and it is widely recognized that acute lung injury (ALI) is often caused by oscillations of cytosolic free Ca2+. Previous studies have indicated that the activation of transient receptor potential-vanilloid (TRPV) channels and subsequent Ca2+ entry initiates an acute calcium-dependent permeability increase during ALI. However, whether seawater exposure induces such an effect through the activation of TRPV channels remains unknown. In the current study, the effect of calcium, a component of seawater, on the inflammatory reactions that occur during seawater drowning-induced ALI, was examined. The results demonstrated that a high concentration of calcium ions in seawater increased lung tissue myeloperoxidase activity and the secretion of inflammatory mediators, such as tumor necrosis factor-α (TNF-α) and interleukin (IL)-1β and IL-6. Further study demonstrated that the seawater challenge elevated cytosolic Ca2+ concentration, indicated by [Ca2+]c, by inducing calcium influx from the extracellular medium via TRPV1 channels. The elevated [Ca2+c] may have resulted in the increased release of TNF-α and IL-1β via increased phosphorylation of nuclear factor-κB (NF-κB). It was concluded that a high concentration of calcium in seawater exacerbated lung injury, and TRPV1 channels were notable mediators of the calcium increase initiated by the seawater challenge. Calcium influx through TRPV1 may have led to greater phosphorylation of NF-κB and increased release of TNF-α and IL-1β. PMID:26796050

  16. Polarized-light interferometry of calcium carbonate deposition in moss from a waterfall on the niagara escarpment.

    PubMed

    Swatland, Howard J

    2010-06-01

    Deposition of calcium carbonate from groundwater was examined on a moss, Didymodon tophaceus, from a Niagara Escarpment waterfall. A spectrophotometer on a polarizing microscope was used for interferometry. A second-order blue interference with an interference minimum around 620 nm was found when moss cell spaces were fully calcified. Filled cell spaces were often surrounded by empty cell spaces. Complete calcification of whole leaflets resulted in progressively higher orders of interference colors and a positive shift in interference minima. Calcified leaflets finally became cemented together, but each retained a weak extinction when rotated. Small calcareous spherulites (mean diameter 15.7 +/- 2.1 microm) were found between leaflets. Spherulites exhibited first-order white interference with a Maltese cross that rotated when the polarizer and analyzer were rotated in tandem. A Nikitin-Berek compensator was tilted at 5.5 degrees to give an interference minimum at 600 nm in the optical axis. Quadrants of spherulites with radii more or less in line with the tilting axis of the compensator had lower (P < 0.001) interference minima (535 +/- 27 nm) than quadrants with radii more or less perpendicular to the compensator (659 +/- 15 nm), thus indicating a radial internal structure. Spherulites were tentatively identified as vaterite.

  17. Calmodulin modulates the delay period between release of calcium from internal stores and activation of calcium influx via endogenous TRP1 channels.

    PubMed

    Vaca, Luis; Sampieri, Alicia

    2002-11-01

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

  18. Large-conductance calcium-activated potassium current modulates excitability in isolated canine intracardiac neurons

    PubMed Central

    Pérez, Guillermo J.; Desai, Mayurika; Anderson, Seth

    2013-01-01

    We studied principal neurons from canine intracardiac (IC) ganglia to determine whether large-conductance calcium-activated potassium (BK) channels play a role in their excitability. We performed whole cell recordings in voltage- and current-clamp modes to measure ion currents and changes in membrane potential from isolated canine IC neurons. Whole cell currents from these neurons showed fast- and slow-activated outward components. Both current components decreased in the absence of calcium and following 1–2 mM tetraethylammonium (TEA) or paxilline. These results suggest that BK channels underlie these current components. Single-channel analysis showed that BK channels from IC neurons do not inactivate in a time-dependent manner, suggesting that the dynamic of the decay of the fast current component is akin to that of intracellular calcium. Immunohistochemical studies showed that BK channels and type 2 ryanodine receptors are coexpressed in IC principal neurons. We tested whether BK current activation in these neurons occurred via a calcium-induced calcium release mechanism. We found that the outward currents of these neurons were not affected by the calcium depletion of intracellular stores with 10 mM caffeine and 10 μM cyclopiazonic acid. Thus, in canine intracardiac neurons, BK currents are directly activated by calcium influx. Membrane potential changes elicited by long (400 ms) current injections showed a tonic firing response that was decreased by TEA or paxilline. These data strongly suggest that the BK current present in canine intracardiac neurons regulates action potential activity and could increase these neurons excitability. PMID:23195072

  19. Effect of polyphenols on calcium content and alkaline phosphatase activity in rat femoral tissues in vitro.

    PubMed

    Yamaguchi, M; Jie, Z

    2001-12-01

    The effect of various polyphenols on calcium content and alkaline phosphatase activity in the femoral-diaphyseal and -metaphyseal tissues of young rats in vitro was investigated. Bone tissues were cultured for 24 h in serum-free Dulbecco's modified Eagle's medium containing either vehicle or various polyphenols (10(-7) - 10(-4) M). The presence of genistein (10(-6) - 10(-4) M) caused a significant increase in calcium content and alkaline phosphatase activity in the femoral-diaphyseal and -metaphyseal tissues. Resveratrol (10(-4) m) decreased metaphyseal calcium content significantly, and it (10(-6) - 10(-4) M) had a significant inhibitory effect on diaphyseal enzyme activity. Epigallocatechin gallate (EGCg; 10(-4) M) significantly inhibited alkaline phosphatase activity in the diaphyseal and metaphyseal tissues. EGCg (10(-7) - 10(-4) M) had no effect on bone calcium content. Meanwhile, glycitein, quercetin, or catechin in the range of 10(-7) to 10(-4) ml did not have an effect on calcium content and alkaline phosphatase activity in the femoral-diaphyseal and -metaphyseal tissues. The present study suggests that a phytoestrogen genistein has a unique anabolic effect on bone calcification in vitro.

  20. Activation of the MAP Kinase Cascade by Exogenous Calcium-Sensing Receptor

    SciTech Connect

    Hobson, Susan A.; Wright, Jay W.; Lee, Fred; Mcneil, Scott; Bilderback, Tim R.; Rodland, Karin D.

    2003-02-01

    In Rat-1 fibroblasts and ovarian surface epithelial cells, extracellular calcium induces a proliferative response which appears to be mediated by the G-protein coupled Calcium-sensing Receptor (CaR), as expression of the non-functional CaR-R795W mutant inhibits both thymidine incorporation and activation of the extracellular-regulated kinase (ERK) in response to calcium. In this report we utilized CaR-transfected HEK293 cells to demonstrate that functional CaR is necessary and sufficient for calcium-induced ERK activation. CaR-dependent ERK activation was blocked by co-expression of the Ras dominant-negative mutant, Ras N17, and by exposure to the phosphatidyl inositol 3' kinase inhibitors wortmannin and LY294002. In contrast to Rat-1 fibroblasts, CaR-mediated in vitro kinase activity of ERK2 was unaffected by tyrosine kinase inhibitor herbimycin in CaR-transfected HEK293 cells. These results suggest that usage of distinct pathways downstream of the CaR varies in a cell-type specific manner, suggesting a potential mechanism by which activation of the CaR could couple to distinct calcium-dependent responses.

  1. Studies of a calcium-activated neutral protease from chicken skeletal muscle. I. Purification and characterization.

    PubMed

    Ishiura, S; Murofushi, H; Suzuki, K; Imahori, K

    1978-07-01

    A calcium-activated neutral protease was purified 2,700-fold over the crude extract from chicken skeletal muscle. The purified protease migrated as a single band on polyacrylamide gel electrophoresis with or without SDS. Its molecular weight was 80,000 and pH optimum for activity was 7.7. The activity required strictly the presence of calcium (optimum concentration: 1.8 mM) or strontium (optimum concentration: 10 mM) ions. The protease was inhibited by leupeptin, which is known to be a strong inhibitor of papain, cathepsin B, trypsin, and plasmin.

  2. A free calcium wave traverses the activating egg of the medaka, Oryzias latipes

    PubMed Central

    Gilkey, J. C.; Jaffe, L. F.; Ridgway, E. B.; Reynolds, G. T.

    1978-01-01

    Aequorin-injected eggs of the medaka (a fresh water fish) show an explosive rise in free calcium during fertilization, which is followed by a slow return to the resting level. Image intensification techniques now show a spreading wave of high free calcium during fertilization. The wave starts at the animal pole (where the sperm enters) and then traverses the egg as a shallow, roughly 20 degrees-wide band which vanishes at the antipode some minutes later. The peak free calcium concentration within this moving band is estimated to be about 30 microM (perhaps 100-1,000 times the resting level). Eggs activated by ionophore A23187 may show multiple initiation sites. The resulting multiple waves never spread through each other; rather, they fuse upon meeting so as to form spreading waves of compound origin. The fertilization wave is nearly independent of extracellular calcium because it is only slightly slowed (by perhaps 15%) in a medium containing 5 mM ethylene glycol-bis[beta-aminoethyl ether]N,N'-tetraacetic acid (EGTA) and no deliberately added calcium. It is also independent of the large cortical vesicles, which may be centrifugally displaced. Normally, however, it distinctly precedes the well-known wave of cortical vesicle exocytosis. We conclude that the fertilization wave in the medaka egg is propagated by calcium-stimulated calcium release, primarily from some internal sources other than the large cortical vesicles. A comparison of the characteristics of the exocytotic wave in the medaka with that in other eggs, particularly in echinoderm eggs, suggests that such a propagated calcium wave is a general feature of egg activation. PMID:10605450

  3. Osteogenic Differentiation of MSC through Calcium Signaling Activation: Transcriptomics and Functional Analysis

    PubMed Central

    Viti, Federica; Landini, Martina; Mezzelani, Alessandra; Petecchia, Loredana; Milanesi, Luciano; Scaglione, Silvia

    2016-01-01

    The culture of progenitor mesenchymal stem cells (MSC) onto osteoconductive materials to induce a proper osteogenic differentiation and mineralized matrix regeneration represents a promising and widely diffused experimental approach for tissue-engineering (TE) applications in orthopaedics. Among modern biomaterials, calcium phosphates represent the best bone substitutes, due to their chemical features emulating the mineral phase of bone tissue. Although many studies on stem cells differentiation mechanisms have been performed involving calcium-based scaffolds, results often focus on highlighting production of in vitro bone matrix markers and in vivo tissue ingrowth, while information related to the biomolecular mechanisms involved in the early cellular calcium-mediated differentiation is not well elucidated yet. Genetic programs for osteogenesis have been just partially deciphered, and the description of the different molecules and pathways operative in these differentiations is far from complete, as well as the activity of calcium in this process. The present work aims to shed light on the involvement of extracellular calcium in MSC differentiation: a better understanding of the early stage osteogenic differentiation program of MSC seeded on calcium-based biomaterials is required in order to develop optimal strategies to promote osteogenesis through the use of new generation osteoconductive scaffolds. A wide spectrum of analysis has been performed on time-dependent series: gene expression profiles are obtained from samples (MSC seeded on calcium-based scaffolds), together with related microRNAs expression and in vivo functional validation. On this basis, and relying on literature knowledge, hypotheses are made on the biomolecular players activated by the biomaterial calcium-phosphate component. Interestingly, a key role of miR-138 was highlighted, whose inhibition markedly increases osteogenic differentiation in vitro and enhance ectopic bone formation in vivo

  4. NMDA receptor-mediated epileptiform persistent activity requires calcium release from intracellular stores in prefrontal neurons.

    PubMed

    Gao, Wen-Jun; Goldman-Rakic, Patricia S

    2006-02-01

    Various normal and pathological forms of synchronized population activity are generated by recurrent excitation among pyramidal neurons in the neocortex. However, the intracellular signaling mechanisms underlying this activity remain poorly understood. In this study, we have examined the cellular properties of synchronized epileptiform activity in the prefrontal cortex with particular emphasis on a potential role of intracellular calcium stores. We find that the zero-magnesium-induced synchronized activity is blocked by inhibition of sarco-endoplasmic reticulum Ca(2+)-ATPases, phospholipase C (PLC), the inositol 1,4,5-trisphosphate (IP3) receptor, and the ryanodine receptor. This same activity is, however, not affected by application of metabotropic glutamatergic receptor (mGluR) agonists, nor by introduction of an mGluR antagonist. These results suggest that persistent synchronized activity in vitro is dependent upon calcium release from internal calcium stores through the activation of PLC-IP3 receptor pathway. Our findings also raise the possibility that intracellular calcium release may be involved in the generation of pathologic synchronized activity in epilepsy in vivo and in physiological forms of synchronized cortical activity.

  5. Activation of the exchange factor Ras-GRF by calcium requires an intact Dbl homology domain.

    PubMed

    Freshney, N W; Goonesekera, S D; Feig, L A

    1997-04-21

    Ras-GRF is a guanine nucleotide exchange factor that activates Ras proteins. Its activity on Ras in cells is enhanced upon calcium influx. Activation follows calcium-induced binding of calmodulin to an IQ motif near the N-terminus of Ras-GRF. Ras-GRF also contains a Dbl homology (DH) domain C-terminal to the IQ motif. In many proteins, DH domains act as exchange factors for Rho-GTPase family members. However, we failed to detect exchange activity of this domain on well characterized Rho family members. Instead, we found that mutations analogous to those that block exchange activity of Dbl prevented Ras-GRF activation by calcium/ calmodulin in vivo. All DH domains are followed immediately by a pleckstrin homology (PH) domain. We found that a mutation at a conserved site within the PH domain following the DH domain also prevented Ras-GRF activation by calcium in vivo. These results suggest that in addition to playing a role as activators of Rho proteins, DH domains can also contribute to the coupling of cellular signals to Ras activation.

  6. Nitric Oxide Regulates Neuronal Activity via Calcium-Activated Potassium Channels

    PubMed Central

    Zhong, Lei Ray; Estes, Stephen; Artinian, Liana; Rehder, Vincent

    2013-01-01

    Nitric oxide (NO) is an unconventional membrane-permeable messenger molecule that has been shown to play various roles in the nervous system. How NO modulates ion channels to affect neuronal functions is not well understood. In gastropods, NO has been implicated in regulating the feeding motor program. The buccal motoneuron, B19, of the freshwater pond snail Helisoma trivolvis is active during the hyper-retraction phase of the feeding motor program and is located in the vicinity of NO-producing neurons in the buccal ganglion. Here, we asked whether B19 neurons might serve as direct targets of NO signaling. Previous work established NO as a key regulator of growth cone motility and neuronal excitability in another buccal neuron involved in feeding, the B5 neuron. This raised the question whether NO might modulate the electrical activity and neuronal excitability of B19 neurons as well, and if so whether NO acted on the same or a different set of ion channels in both neurons. To study specific responses of NO on B19 neurons and to eliminate indirect effects contributed by other cells, the majority of experiments were performed on single cultured B19 neurons. Addition of NO donors caused a prolonged depolarization of the membrane potential and an increase in neuronal excitability. The effects of NO could mainly be attributed to the inhibition of two types of calcium-activated potassium channels, apamin-sensitive and iberiotoxin-sensitive potassium channels. NO was found to also cause a depolarization in B19 neurons in situ, but only after NO synthase activity in buccal ganglia had been blocked. The results suggest that NO acts as a critical modulator of neuronal excitability in B19 neurons, and that calcium-activated potassium channels may serve as a common target of NO in neurons. PMID:24236040

  7. 12 CFR 347.206 - Domestic retail deposit activity requiring deposit insurance by U.S. branch of a foreign bank.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 12 Banks and Banking 4 2010-01-01 2010-01-01 false Domestic retail deposit activity requiring... Banks § 347.206 Domestic retail deposit activity requiring deposit insurance by U.S. branch of a foreign bank. (a) Domestic retail deposit activity. To initiate or conduct domestic retail deposit...

  8. Agonist-Biased Signaling via Proteinase Activated Receptor-2: Differential Activation of Calcium and Mitogen-Activated Protein Kinase Pathways

    PubMed Central

    Ramachandran, Rithwik; Mihara, Koichiro; Mathur, Maneesh; Rochdi, Moulay Driss; Bouvier, Michel; DeFea, Kathryn

    2009-01-01

    We evaluated the ability of different trypsin-revealed tethered ligand (TL) sequences of rat proteinase-activated receptor 2 (rPAR2) and the corresponding soluble TL-derived agonist peptides to trigger agonist-biased signaling. To do so, we mutated the proteolytically revealed TL sequence of rPAR2 and examined the impact on stimulating intracellular calcium transients and mitogen-activated protein (MAP) kinase. The TL receptor mutants, rPAR2-Leu37Ser38, rPAR2-Ala37–38, and rPAR2-Ala39–42 were compared with the trypsin-revealed wild-type rPAR2 TL sequence, S37LIGRL42—. Upon trypsin activation, all constructs stimulated MAP kinase signaling, but only the wt-rPAR2 and rPAR2-Ala39–42 triggered calcium signaling. Furthermore, the TL-derived synthetic peptide SLAAAA-NH2 failed to cause PAR2-mediated calcium signaling but did activate MAP kinase, whereas SLIGRL-NH2 triggered both calcium and MAP kinase signaling by all receptors. The peptides AAIGRL-NH2 and LSIGRL-NH2 triggered neither calcium nor MAP kinase signals. Neither rPAR2-Ala37–38 nor rPAR2-Leu37Ser38 constructs recruited β-arrestins-1 or -2 in response to trypsin stimulation, whereas both β-arrestins were recruited to these mutants by SLIGRL-NH2. The lack of trypsin-triggered β-arrestin interactions correlated with impaired trypsin-activated TL-mutant receptor internalization. Trypsin-stimulated MAP kinase activation by the TL-mutated receptors was not blocked by inhibitors of Gαi (pertussis toxin), Gαq [N-cyclohexyl-1-(2,4-dichlorophenyl)-1,4-dihydro-6-methylindeno[1,2-c]pyrazole-3-carboxamide (GP2A)], Src kinase [4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]-pyrimidine (PP1)], or the epidermal growth factor (EGF) receptor [4-(3′-chloroanilino)-6,7-dimethoxy-quinazoline (AG1478)], but was inhibited by the Rho-kinase inhibitor (R)-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexanecarboxamide, 2HCl (Y27362). The data indicate that the proteolytically revealed TL sequence(s) and the mode

  9. Ultrastructural characteristics of fibrin clots from canine and feline platelet concentrates activated with calcium gluconate or calcium gluconate plus batroxobin

    PubMed Central

    2013-01-01

    Background The aim of this study was to use transmission electron microscopy to describe the ultrastructural characteristics of clots obtained from canine and feline platelet concentrates (PC) that had been activated with calcium gluconate (CG) or CG plus batroxobin (CGB). Platelets from fibrin clots were classified according their morphological changes. The area of the intercellular space (μm2), the area of the fibrin fibers (μm2), and the width of the fibrin fibers (μm) were determined for the dog clots. The platelet area (μm2), the area of fibrin fibers (μm2), the ratio of the minor and major axes of platelets, the ratio of the major and minor axes of platelets, and the number of α-granules found within platelets were measured for the cat clots. Results Cat platelets displayed full activation. Dog platelets displayed lysis with loss of normal architecture. In both species, a statistically significant difference was found (P < 0.01) between the fibrin fiber measurements in the PC clots activated with CG and CGB. Conclusions The findings suggest that activation with CG caused platelet alpha granules to release their contents. In cats, fibrin production was greater when the PC was activated with CG. In dogs, activation with CG produced thick fibrin fibers. PMID:23587176

  10. The selective activation of the cardiac sarcolemmal sodium-calcium exchanger by plasmalogenic phosphatidic acid produced by phospholipase D.

    PubMed

    Hale, C C; Ebeling, E G; Hsu, F F; Ford, D A

    1998-01-30

    Since plasmalogens are the predominant phospholipid of cardiac sarcolemma, the activation of the sodium-calcium exchanger by either plasmenylethanolamine or plasmalogenic phosphatidic acid generated by phospholipase D was explored. Sodium-calcium exchange activity was 7-fold greater in proteoliposomes comprised of plasmenylethanolamine compared to proteoliposomes comprised of only plasmenylcholine. Phospholipase D treatment of proteoliposomes resulted in 1 mol % conversion of plasmenylcholine or phosphatidylcholine to their respective phosphatidic acid molecular species with a concomitant 8-fold or 2-fold activation of sodium-calcium exchange activity, respectfully. Thus, phospholipase D-mediated hydrolysis of plasmalogens to phosphatidic acid may be an important mechanism for the regulation of the sodium-calcium exchanger.

  11. Localization of intracellular calcium release in cells injured by venom from the ectoparasitoid Nasonia vitripennis (Walker) (Hymenoptera: Pteromalidae) and dependence of calcium mobilization on G-protein activation.

    PubMed

    Rivers, David B; Crawley, Timothy; Bauser, Holly

    2005-02-01

    Venom from the ectoparasitic wasp Nasonia vitripennis induces cellular injury that appears to involve the release of intracellular calcium stores via the activation of phospholipase C, and culminates in oncotic death. A linkage between release of intracellular Ca2+ and oncosis has not been clearly established and was the focus of this study. When BTI-TN-5B1-4 cells were treated with suramin, an uncoupler of G-proteins, venom-induced swelling and oncotic death were inhibited in a dose-dependent manner for at least 24 h. Suramin also blocked increases in free cytosolic [Ca2+], arguing that venom induces calcium mobilization through G-protein signaling pathways. Endoplasmic reticulum (ER) was predicted to be the source of intracellular calcium release, but labeling with the fluorescent probe ER-tracker revealed no indication of organelle swelling or loss of membrane integrity as would be expected if the Ca(2+)-ATPase pump was disabled by crude venom. Incubation of cell monolayers with calmodulin or nitrendipine, modulators of ER calcium release channels, neither attenuated nor augmented the effects of wasp venom. These results suggest that wasp venom stimulates calcium release from ER compartments distinct from RyRs, L-type Ca2+ channels, and the Ca(2+)-ATPase pump, or calcium is released from some other intracellular store. A reduction of mitochondrial membrane potential delta psi(m) appeared to precede a rise in cytosolic free Ca2+ as evidenced by fluorescent microscopy using the calcium-sensitive probe fluo-4 AM. This argues that the initial insult to the cell resulting from venom elicits a rapid loss of (delta psi(m)), followed by unregulated calcium efflux from mitochondria into the cytosol. Mobilization of calcium in this fashion could stimulate cAMP formation, and subsequently promote calcium release from NAADP-sensitive stores.

  12. The effect of crystal structure of TiO2 nanotubes on the formation of calcium phosphate coatings during biomimetic deposition

    NASA Astrophysics Data System (ADS)

    Liu, Yi; Kim, Sun; McLeod, John A.; Li, Jun; Guo, Xiaoxuan; Sham, Tsun-Kong; Liu, Lijia

    2017-02-01

    The crystallization process of bioactive calcium phosphate (CaP) species via biomimetic deposition onto anodic TiO2 nanotubes is investigated. The porous surface of nanostructured TiO2 provides an ideal substrate for CaP crystallization. The compositions of CaP coatings are studied using X-ray absorption near-edge structures (XANES) at the Ca K-edge. Using detection modes with different probing depths, both the surface of the CaP coating and the CaP-TiO2 interface are simultaneously analyzed. Calcium phosphate (CaP) species, such as hydroxyapatite (HAp), octacalcium phosphate (Ca8(HPO4)2(PO4)4·5H2O, OCP), brushite (CaHPO4·2H2O, DCPD), and amorphous calcium phosphate (ACP), are found in the CaP coatings. TiO2 nanotubes of amorphous and anatase phases are comparatively studied to determine their effect on the efficiency of CaP formation and the phase transformation among CaP species in prolonged deposition time. It is found the composition of CaP coating has a strong dependency on the crystal structure of TiO2 substrate and the kinetics (deposition time).

  13. Familial Alzheimer’s disease–associated presenilin-1 alters cerebellar activity and calcium homeostasis

    PubMed Central

    Sepulveda-Falla, Diego; Barrera-Ocampo, Alvaro; Hagel, Christian; Korwitz, Anne; Vinueza-Veloz, Maria Fernanda; Zhou, Kuikui; Schonewille, Martijn; Zhou, Haibo; Velazquez-Perez, Luis; Rodriguez-Labrada, Roberto; Villegas, Andres; Ferrer, Isidro; Lopera, Francisco; Langer, Thomas; De Zeeuw, Chris I.; Glatzel, Markus

    2014-01-01

    Familial Alzheimer’s disease (FAD) is characterized by autosomal dominant heritability and early disease onset. Mutations in the gene encoding presenilin-1 (PS1) are found in approximately 80% of cases of FAD, with some of these patients presenting cerebellar damage with amyloid plaques and ataxia with unclear pathophysiology. A Colombian kindred carrying the PS1-E280A mutation is the largest known cohort of PS1-FAD patients. Here, we investigated PS1-E280A–associated cerebellar dysfunction and found that it occurs early in PS1-E208A carriers, while cerebellar signs are highly prevalent in patients with dementia. Postmortem analysis of cerebella of PS1-E280A carrier revealed greater Purkinje cell (PC) loss and more abnormal mitochondria compared with controls. In PS1-E280A tissue, ER/mitochondria tethering was impaired, Ca2+ channels IP3Rs and CACNA1A were downregulated, and Ca2+-dependent mitochondrial transport proteins MIRO1 and KIF5C were reduced. Accordingly, expression of PS1-E280A in a neuronal cell line altered ER/mitochondria tethering and transport compared with that in cells expressing wild-type PS1. In a murine model of PS1-FAD, animals exhibited mild ataxia and reduced PC simple spike activity prior to cerebellar β-amyloid deposition. Our data suggest that impaired calcium homeostasis and mitochondrial dysfunction in PS1-FAD PCs reduces their activity and contributes to motor coordination deficits prior to Aβ aggregation and dementia. We propose that PS1-E280A affects both Ca2+ homeostasis and Aβ precursor processing, leading to FAD and neurodegeneration. PMID:24569455

  14. Lymphocytes prime activation is required for nicotine-induced calcium waves.

    PubMed

    Landais, Emilie; Liautaud-Roger, Francoise; Antonicelli, Frank

    2010-06-01

    Lymphocytes are reported to express nicotinic acetylcholine receptors (nAChR). However, no data are available on the expression of these nAChR on activated lymphocyte relatively to resting lymphocytes. In this study, we examined nAChR subunits expression in PHA-stimulated versus un-stimulated lymphocytes, and four leukemic cell lines. Cell stimulation with nicotine triggered calcium responses only in some experiments conducted with PHA-stimulated lymphocytes. Likewise, only the Jurkat and HL-60 cell lines displayed calcium waves upon nicotine stimulation, whereas the Raji and CCRF-CEM did not. All responding cells displayed an active form of the nicotinic a-7 nAChR. Indeed, use of 2 different sets of primers for the corresponding mRNA showed that expression of the full-length a-7 subunit mRNA was only present in PHA-stimulated lymphocytes for which calcium waves had been evidenced. Microscopy analysis of lymphocytes structure showed a direct relationship between their size, their a-7 nAChR expression, and calcium release upon nicotine stimulation. Then, this relationship suggested that lymphocytes need a prime activation to express the a-7 nAChR, and therefore to release calcium in response to nicotine.

  15. Activity-dependent regulation of T-type calcium channels by submembrane calcium ions

    PubMed Central

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

    2017-01-01

    Voltage-gated Ca2+ channels are involved in numerous physiological functions and various mechanisms finely tune their activity, including the Ca2+ ion itself. This is well exemplified by the Ca2+-dependent inactivation of L-type Ca2+ channels, whose alteration contributes to the dramatic disease Timothy Syndrome. For T-type Ca2+ channels, a long-held view is that they are not regulated by intracellular Ca2+. Here we challenge this notion by using dedicated electrophysiological protocols on both native and expressed T-type Ca2+ channels. We demonstrate that a rise in submembrane Ca2+ induces a large decrease in T-type current amplitude due to a hyperpolarizing shift in the steady-state inactivation. Activation of most representative Ca2+-permeable ionotropic receptors similarly regulate T-type current properties. Altogether, our data clearly establish that Ca2+ entry exerts a feedback control on T-type channel activity, by modulating the channel availability, a mechanism that critically links cellular properties of T-type Ca2+ channels to their physiological roles. DOI: http://dx.doi.org/10.7554/eLife.22331.001 PMID:28109159

  16. Activity-dependent regulation of T-type calcium channels by submembrane calcium ions.

    PubMed

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

    2017-01-21

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

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

    PubMed

    Goldberg, Joshua A; Wilson, Charles J

    2005-11-02

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

  18. Sigma-1 receptor stimulation attenuates calcium influx through activated L-type Voltage Gated Calcium Channels in purified retinal ganglion cells.

    PubMed

    Mueller, Brett H; Park, Yong; Daudt, Donald R; Ma, Hai-Ying; Akopova, Irina; Stankowska, Dorota L; Clark, Abbot F; Yorio, Thomas

    2013-02-01

    Sigma-1 receptors (σ-1rs) exert neuroprotective effects on retinal ganglion cells (RGCs) both in vivo and in vitro. This receptor has unique properties through its actions on several voltage-gated and ligand-gated channels. The purpose of this study was to investigate the role that σ-1rs play in regulating cell calcium dynamics through activated L-type Voltage Gated Calcium Channels (L-type VGCCs) in purified RGCs. RGCs were isolated from P3-P7 Sprague-Dawley rats and purified by sequential immunopanning using a Thy1.1 antibody. Calcium imaging was used to measure changes in intracellular calcium after depolarizing the cells with potassium chloride (KCl) in the presence or absence of two σ-1r agonists [(+)-SKF10047 and (+)-Pentazocine], one σ-1r antagonist (BD1047), and one L-type VGCC antagonist (Verapamil). Finally, co-localization studies were completed to assess the proximity of σ-1r with L-type VGCCs in purified RGCs. VGCCs were activated using KCl (20 mM). Pre-treatment with a known L-type VGCC blocker demonstrated a 57% decrease of calcium ion influx through activated VGCCs. Calcium imaging results also demonstrated that σ-1r agonists, (+)-N-allylnormetazocine hydrochloride [(+)-SKF10047] and (+)-Pentazocine, inhibited calcium ion influx through activated VGCCs. Antagonist treatment using BD1047 demonstrated a potentiation of calcium ion influx through activated VGCCs and abolished all inhibitory effects of the σ-1r agonists on VGCCs, implying that these ligands were acting through the σ-1r. An L-type VGCC blocker (Verapamil) also inhibited KCl activated VGCCs and when combined with the σ-1r agonists there was not a further decline in calcium entry suggesting similar mechanisms. Lastly, co-localization studies demonstrated that σ-1rs and L-type VGCCs are co-localized in purified RGCs. Taken together, these results indicated that σ-1r agonists can inhibit KCl induced calcium ion influx through activated L-type VGCCs in purified RGCs. This is the

  19. Calcium signaling and the MAPK cascade are required for sperm activation in Caenorhabditis elegans.

    PubMed

    Liu, Zhiyu; Wang, Bin; He, Ruijun; Zhao, Yanmei; Miao, Long

    2014-02-01

    In nematode, sperm activation (or spermiogenesis), a process in which the symmetric and non-motile spermatids transform into polarized and crawling spermatozoa, is critical for sperm cells to acquire fertilizing competence. SPE-8 dependent and SPE-8 independent pathways function redundantly during sperm activation in both males and hermaphrodites of Caenorhabditis elegans. However, the downstream signaling for both pathways remains unclear. Here we show that calcium signaling and the MAPK cascade are required for both SPE-8 dependent and SPE-8 independent sperm activation, implying that both pathways share common downstream signaling components during sperm activation. We demonstrate that activation of the MAPK cascade is sufficient to activate spermatids derived from either wild-type or spe-8 group mutant males and that activation of the MAPK cascade bypasses the requirement of calcium signal to induce sperm activation, indicating that the MAPK cascade functions downstream of or parallel with the calcium signaling during sperm activation. Interestingly, the persistent activation of MAPK in activated spermatozoa inhibits Major Sperm Protein (MSP)-based cytoskeleton dynamics. We demonstrate that MAPK plays dual roles in promoting pseudopod extension during sperm activation but also blocking the MSP-based, amoeboid motility of the spermatozoa. Thus, though nematode sperm are crawling cells, morphologically distinct from flagellated sperm, and the molecular machinery for motility of amoeboid and flagellated sperm is different, both types of sperm might utilize conserved signaling pathways to modulate sperm maturation.

  20. Deposition of Carbonated Hydroxyapatite (CO3HAp) on Poly(Methylmethacrylate Surfaces by Decomposition of Calcium-EDTA Chelate

    SciTech Connect

    Yusufoglu, Yusuf; Akinc, Mufit

    2008-10-01

    Inspired from the nature, the development of organic-inorganic composites between polymers and hydroxyapatite (HAp) has been investigated extensively. In this study, bone-like apatite (carbonated hydroxyapatite, CO{sub 3}HAp) was precipitated on poly(methylmethacrylate) (PMMA) films by the oxidative decomposition of Ca-EDTA (calcium-ethylenediamine tetraacetate) chelates. Corona-treated PMMA films were soaked in a Ca-EDTA-PO{sub 4}-H{sub 2}O{sub 2} solution and aged at 63 C and pH-9 for times ranging from 1 to 24 h. Apatite formed on PMMA films was characterized by X-ray diffraction, Fourier transform infrared (FTIR), Scanning electron microscope, energy-dispersive X-ray spectroscopy, and carbon analysis. The apatite was found to be CO{sub 3}HAp with Ca/P atomic ratio ranging between 1.3 and 1.9. Elemental analyses indicated that the carbonate content of the apatite phase was around 6.5 wt% after 24 h of aging time. Lattice parameters were estimated using a Rietveld profile-analysis and found to be a = 0.9438 nm and c = 0.6901 nm. Furthermore, FTIR spectra indicated that the apatite deposited on PMMA was B-type CO{sub 3}HAp, in which carbonate ions occupy the phosphate sites. In the first 3 h of aging, isolated rod-like HAp particles were observed. With time, the needle-like crystallites radiate from a nucleus to form double-spherulite shape particles. The crystallites grew into a continuous layer with a thickness of -15 {micro}m after 24-h aging. The adhesive strength between the PMMA substrate and the apatite layer was determined to be around 1.7 MPa.

  1. Direct deposited porous scaffolds of calcium phosphate cement with alginate for drug delivery and bone tissue engineering.

    PubMed

    Lee, Gil-Su; Park, Jeong-Hui; Shin, Ueon Sang; Kim, Hae-Won

    2011-08-01

    This study reports the preparation of novel porous scaffolds of calcium phosphate cement (CPC) combined with alginate, and their potential usefulness as a three-dimensional (3-D) matrix for drug delivery and tissue engineering of bone. An α-tricalcium phosphate-based powder was mixed with sodium alginate solution and then directly injected into a fibrous structure in a Ca-containing bath. A rapid hardening reaction of the alginate with Ca(2+) helps to shape the composite into a fibrous form with diameters of hundreds of micrometers, and subsequent pressing in a mold allows the formation of 3-D porous scaffolds with different porosity levels. After transformation of the CPC into a calcium-deficient hydroxyapatite phase in simulated biological fluid the scaffold was shown to retain its mechanical stability. During the process biological proteins, such as bovine serum albumin and lysozyme, used as model proteins, were observed to be effectively loaded onto and released from the scaffolds for up to more than a month, proving the efficacy of the scaffolds as a drug delivering matrix. Mesenchymal stem cells (MSCs) were isolated from rat bone marrow and then cultured on the CPC-alginate porous scaffolds to investigate the ability to support proliferation of cells and their subsequent differentiation along the osteogenic lineage. It was shown that MSCs increasingly actively populated and also permeated into the porous network with time of culture. In particular, cells cultured within a scaffold with a relatively high porosity level showed favorable proliferation and osteogenic differentiation. An in vivo pilot study of the CPC-alginate porous scaffolds after implantation into the rat calvarium for 6 weeks revealed the formation of new bone tissue within the scaffold, closing the defect almost completely. Based on these results, the newly developed CPC-alginate porous scaffolds could be potentially useful as a 3-D matrix for drug delivery and tissue engineering of bone.

  2. Enhanced Bactericidal Activity of Silver Thin Films Deposited via Aerosol-Assisted Chemical Vapor Deposition.

    PubMed

    Ponja, Sapna D; Sehmi, Sandeep K; Allan, Elaine; MacRobert, Alexander J; Parkin, Ivan P; Carmalt, Claire J

    2015-12-30

    Silver thin films were deposited on SiO2-barrier-coated float glass, fluorine-doped tin oxide (FTO) glass, Activ glass, and TiO2-coated float glass via AACVD using silver nitrate at 350 °C. The films were annealed at 600 °C and analyzed by X-ray powder diffraction, X-ray photoelectron spectroscopy, UV/vis/near-IR spectroscopy, and scanning electron microscopy. All the films were crystalline, and the silver was present in its elemental form and of nanometer dimension. The antibacterial activity of these samples was tested against Escherichia coli and Staphylococcus aureus in the dark and under UV light (365 nm). All Ag-deposited films reduced the numbers of E. coli by 99.9% within 6 h and the numbers of S. aureus by 99.9% within only 2 h. FTO/Ag reduced bacterial numbers of E. coli to below the detection limit after 60 min and caused a 99.9% reduction of S. aureus within only 15 min of UV irradiation. Activ/Ag reduced the numbers of S. aureus by 66.6% after 60 min and TiO2/Ag killed 99.9% of S. aureus within 60 min of UV exposure. More remarkably, we observed a 99.9% reduction in the numbers of E. coli within 6 h and the numbers of S. aureus within 4 h in the dark using our novel TiO2/Ag system.

  3. Increased intracellular calcium activates serum and glucocorticoid-inducible kinase 1 (SGK1) through a calmodulin-calcium calmodulin dependent kinase kinase pathway in Chinese hamster ovary cells.

    PubMed

    Imai, Seiji; Okayama, Naotsuka; Shimizu, Manabu; Itoh, Makoto

    2003-04-04

    SGK1 is one of the protein-serine/threonine kinases that is activated by insulin in a PI3K-dependent manner. Although SGK1 mediates a variety of biological activities, the mechanisms regulating its activity remain unclear. In this study, we examined the potential roles of calcium signaling in the activation of SGK1. Treatment of CHO-IR cells with a cell-permeable calcium chelator, BAPTA-AM, abolished the insulin-induced activation of SGK1. Increasing intracellular calcium concentration by treating cells with thapsigargin or ionomycin induced a 6-8 fold increase in SGK1 activation. This was not affected by a PI3K inhibitor, wortmannin, but was completely inhibited by the calmodulin inhibitors, W 7 and W 5. Co-transfection of CHO cells with FLAG-SGK1 and CaMKK revealed the direct association of CaMKK with SGK1. These results suggest a calcium-triggered signaling cascade in which an increase in intracellular calcium concentration directly stimulates SGK1 through CaMKK.

  4. Complex activity patterns in arterial wall: results from a model of calcium dynamics.

    PubMed

    Buchner, Teodor; Pietkun, Jakub; Kuklik, Paweł

    2012-03-01

    Using a dynamical model of smooth muscle cells in an arterial wall, defined as a system of coupled five-dimensional nonlinear oscillators, on a grid with cylindrical symmetry, we compare the admissible activity patterns with those known from the heart tissue. We postulate on numerical basis the possibility to induce a stable spiral wave in the arterial wall. Such a spiral wave can inhibit the propagation of the axial calcium wave and effectively stop the vasomotion. We also discuss the dynamics of the circumferential calcium wave in comparison to rotors in venous ostia that are a common source of supraventricular ectopy. We show that the velocity and in consequence the frequency range of the circumferential calcium wave is by orders of magnitude too small compared to that of the rotors. The mechanism of the rotor is not likely to involve the calcium-related dynamics of the smooth muscle cells. The calcium-related dynamics which is voltage-independent and hard to be reset seems to actually protect the blood vessels against the electric activity of the atria. We also discuss the microreentry phenomenon, which was found in numerical experiments in the studied model.

  5. Changes in calpain and calpastatin activities of osmotically dehydrated bovine muscle during storage after treatment with calcium.

    PubMed

    Gerelt, B; Rusman, H; Nishiumi, T; Suzuki, A

    2005-05-01

    Calpain and calpastatin activities were investigated in calcium-treated beef after osmotic dehydration. Dehydrated beef was soaked in 150 mM calcium chloride solution for 3 h, and then stored for 48 h at 3-4 °C. The untreated sample (control) was soaked in deionized water for 3 h instead of calcium chloride solution, after osmotic dehydration. The increase and decrease in the relative activity of crude calpain were observed in the untreated and the calcium-treated meat, respectively, during the storage. When the crude calpains were subjected to DEAE-Sephacel column chromatography, it was found that μ-calpain activity decreased rapidly during the storage in the untreated meat, whereas there was almost no change in the activity of m-calpain during the storage. The decrease of calpastatin activity was moderate compared with the decrease of μ-calpain activity. In the calcium chloride-treated meat, however, no μ-calpain nor calpastatin activities was detectable after 48 h at cold-room temperature, and m-calpain activity after 48 h had decreased to 6.1% of its activity immediately after thawing. It was concluded that 150 mM calcium chloride treatment after osmotic dehydration was sufficient to introduce calcium ions into the meat. In the presence of sufficient calcium, autolysis of calpains and proteolytic degradation of calpastatin, which eventually related to the rate of decrease in calpain and calpastatin activities, clearly seem to be related to a decrease in meat toughness.

  6. Calcium current and calcium-activated inward current in the oocyte of the starfish Leptasterias hexactis.

    PubMed Central

    Lansman, J B

    1987-01-01

    1. Inward currents in the immature oocyte of the starfish Leptasterias hexactis were studied with a two-micro-electrode voltage clamp. Experiments investigated the role of Ca2+ in the Na+-dependent plateau of the action potential. 2. Voltage steps more positive than -55 mV produced inward currents in normal sea water that activated and then decayed to a non-zero level with a double-exponential time course. Returning the voltage to the resting potential produced an inward tail current that relaxed slowly to zero with a time course of seconds. 3. Replacing Na+ with choline abolished the slowly decaying component as well as the slow tail current which followed the end of the voltage pulse. This suggested that inward current in Na+-containing sea water consisted of a rapidly decaying component that flowed through Ca2+ channels and a more slowly decaying component carried by Na+. 4. Ca2+ current was isolated in Na+-free sea water. Activation followed a sigmoidal time course that could be described with m2 kinetics. Inactivation during a maintained depolarization followed first-order kinetics and was voltage dependent. 5. When Ba2+ was substituted for Ca2+ as the divalent ion charge carrier, inward currents in Na+-containing sea water decayed along a single-exponential time course. The absence of a slowly decaying Na+ current in Ba2+-containing sea water suggested that Na+ current depended on Ca2+ influx. 6. The effects of altering Ca2+ influx on the time course of Na+ current were investigated. Na+ current decayed more rapidly as the test pulse potential was made more positive, while raising [Ca2+]o slowed the decaying phase without altering its dependence on membrane potential. 7. Tail currents measured after rapidly stepping the membrane potential back to the resting level consisted of a fast component associated with the closing of Ca2+ channels and a slow component that was abolished by removing Na+. 8. The variation of the amplitude of the slow component of tail

  7. Location of Release Sites and Calcium-Activated Chloride Channels Relative to Calcium Channels at the Photoreceptor Ribbon Synapse

    PubMed Central

    Mercer, A. J.; Rabl, K.; Riccardi, G. E.; Brecha, N. C.; Stella, S. L.

    2011-01-01

    Vesicle release from photoreceptor ribbon synapses is regulated by L-type Ca2+ channels, which are in turn regulated by Cl− moving through calcium-activated chloride [Cl(Ca)] channels. We assessed the proximity of Ca2+ channels to release sites and Cl(Ca) channels in synaptic terminals of salamander photoreceptors by comparing fast (BAPTA) and slow (EGTA) intracellular Ca2+ buffers. BAPTA did not fully block synaptic release, indicating some release sites are <100 nm from Ca2+ channels. Comparing Cl(Ca) currents with predicted Ca2+ diffusion profiles suggested that Cl(Ca) and Ca2+ channels average a few hundred nanometers apart, but the inability of BAPTA to block Cl(Ca) currents completely suggested some channels are much closer together. Diffuse immunolabeling of terminals with an antibody to the putative Cl(Ca) channel TMEM16A supports the idea that Cl(Ca) channels are dispersed throughout the presynaptic terminal, in contrast with clustering of Ca2+ channels near ribbons. Cl(Ca) currents evoked by intracellular calcium ion concentration ([Ca2+]i) elevation through flash photolysis of DM-nitrophen exhibited EC50 values of 556 and 377 nM with Hill slopes of 1.8 and 2.4 in rods and cones, respectively. These relationships were used to estimate average submembrane [Ca2+]i in photoreceptor terminals. Consistent with control of exocytosis by [Ca2+] nanodomains near Ca2+ channels, average submembrane [Ca2+]i remained below the vesicle release threshold (∼400 nM) over much of the physiological voltage range for cones. Positioning Ca2+ channels near release sites may improve fidelity in converting voltage changes to synaptic release. A diffuse distribution of Cl(Ca) channels may allow Ca2+ influx at one site to influence relatively distant Ca2+ channels. PMID:21084687

  8. Molecular Dynamics Simulations of Membrane-Bound STIM1 to Investigate Conformational Changes during STIM1 Activation upon Calcium Release.

    PubMed

    Mukherjee, Sreya; Karolak, Aleksandra; Debant, Marjolaine; Buscaglia, Paul; Renaudineau, Yves; Mignen, Olivier; Guida, Wayne C; Brooks, Wesley H

    2017-02-27

    Calcium is involved in important intracellular processes, such as intracellular signaling from cell membrane receptors to the nucleus. Typically, calcium levels are kept at less than 100 nM in the nucleus and cytosol, but some calcium is stored in the endoplasmic reticulum (ER) lumen for rapid release to activate intracellular calcium-dependent functions. Stromal interacting molecule 1 (STIM1) plays a critical role in early sensing of changes in the ER's calcium level, especially when there is a sudden release of stored calcium from the ER. Inactive STIM1, which has a bound calcium ion, is activated upon ion release. Following activation of STIM1, there is STIM1-assisted initiation of extracellular calcium entry through channels in the cell membrane. This extracellular calcium entering the cell then amplifies intracellular calcium-dependent actions. At the end of the process, ER levels of stored calcium are reestablished. The main focus of this work was to study the conformational changes accompanying homo- or heterodimerization of STIM1. For this purpose, the ER luminal portion of STIM1 (residues 58-236), which includes the sterile alpha motif (SAM) domain plus the calcium-binding EF-hand domains 1 and 2 attached to the STIM1 transmembrane region (TM), was modeled and embedded in a virtual membrane. Next, molecular dynamics simulations were performed to study the conformational changes that take place during STIM1 activation and subsequent protein-protein interactions. Indeed, the simulations revealed exposure of residues in the EF-hand domains, which may be important for dimerization steps. Altogether, understanding conformational changes in STIM1 can help in drug discovery when targeting this key protein in intracellular calcium functions.

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

    PubMed Central

    Rudolph, Stephanie; Hull, Court

    2015-01-01

    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. SIGNIFICANCE STATEMENT 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

  10. Long-Chain Fatty Acids Activate Calcium Channels in Ventricular Myocytes

    NASA Astrophysics Data System (ADS)

    Huang, James Min-Che; Xian, Hu; Bacaner, Marvin

    1992-07-01

    Nonesterified fatty acids accumulate at sites of tissue injury and necrosis. In cardiac tissue the concentrations of oleic acid, arachidonic acid, leukotrienes, and other fatty acids increase greatly during ischemia due to receptor or nonreceptor-mediated activation of phospholipases and/or diminished reacylation. In ischemic myocardium, the time course of increase in fatty acids and tissue calcium closely parallels irreversible cardiac damage. We postulated that fatty acids released from membrane phospholipids may be involved in the increase of intracellular calcium. We report here that low concentrations (3-30 μM) of each long-chain unsaturated (oleic, linoleic, linolenic, and arachidonic) and saturated (palmitic, stearic, and arachidic) fatty acid tested induced multifold increases in voltage-dependent calcium currents (ICa) in cardiac myocytes. In contrast, neither short-chain fatty acids (<12 carbons) or fatty acid esters (oleic and palmitic methyl esters) had any effect on ICa, indicating that activation of calcium channels depended on chain length and required a free carboxyl group. Inhibition of protein kinases C and A, G proteins, eicosanoid production, or nonenzymatic oxidation did not block the fatty acid-induced increase in ICa. Thus, long-chain fatty acids appear to directly activate ICa, possibly by acting at some lipid sites near the channels or directly on the channel protein itself. We suggest that the combined effects of fatty acids released during ischemia on ICa may contribute to ischemia-induced pathogenic events on the heart that involve calcium, such as arrhythmias, conduction disturbances, and myocardial damage due to cytotoxic calcium overload.

  11. Calcium Pyrophosphate Deposition (CPPD)

    MedlinePlus

    ... Fellows Evidence-Based Practice for Academic Researchers Responsible Data Management in Research Career Planning Treatments Patient and Caregiver ... Fellows Evidence-Based Practice for Academic Researchers Responsible Data Management in Research Career Planning Treatments Patient and Caregiver ...

  12. Variation in human cancer cell external phosphatidylserine is regulated by flippase activity and intracellular calcium.

    PubMed

    Vallabhapurapu, Subrahmanya D; Blanco, Víctor M; Sulaiman, Mahaboob K; Vallabhapurapu, Swarajya Lakshmi; Chu, Zhengtao; Franco, Robert S; Qi, Xiaoyang

    2015-10-27

    Viable cancer cells expose elevated levels of phosphatidylserine (PS) on the exoplasmic face of the plasma membrane. However, the mechanisms leading to elevated PS exposure in viable cancer cells have not been defined. We previously showed that externalized PS may be used to monitor, target and kill tumor cells. In addition, PS on tumor cells is recognized by macrophages and has implications in antitumor immunity. Therefore, it is important to understand the molecular details of PS exposure on cancer cells in order to improve therapeutic targeting. Here we explored the mechanisms regulating the surface PS exposure in human cancer cells and found that differential flippase activity and intracellular calcium are the major regulators of surface PS exposure in viable human cancer cells. In general, cancer cell lines with high surface PS exhibited low flippase activity and high intracellular calcium, whereas cancer cells with low surface PS exhibited high flippase activity and low intracellular calcium. High surface PS cancer cells also had higher total cellular PS than low surface PS cells. Together, our results indicate that the amount of external PS in cancer cells is regulated by calcium dependent flippase activity and may also be influenced by total cellular PS.

  13. Activation and inhibition of TMEM16A calcium-activated chloride channels.

    PubMed

    Ni, Yu-Li; Kuan, Ai-Seon; Chen, Tsung-Yu

    2014-01-01

    Calcium-activated chloride channels (CaCC) encoded by family members of transmembrane proteins of unknown function 16 (TMEM16) have recently been intensely studied for functional properties as well as their physiological roles as chloride channels in various tissues. One technical hurdle in studying these channels is the well-known channel rundown that frequently impairs the precision of electrophysiological measurements for the channels. Using experimental protocols that employ fast-solution exchange, we circumvented the problem of channel rundown by normalizing the Ca(2+)-induced current to the maximally-activated current obtained within a time period in which the channel rundown was negligible. We characterized the activation of the TMEM16A-encoded CaCC (also called ANO1) by Ca(2+), Sr(2+), and Ba(2+), and discovered that Mg(2+) competes with Ca(2+) in binding to the divalent-cation binding site without activating the channel. We also studied the permeability of the ANO1 pore for various anions and found that the anion occupancy in the pore-as revealed by the permeability ratios of these anions-appeared to be inversely correlated with the apparent affinity of the ANO1 inhibition by niflumic acid (NFA). On the other hand, the NFA inhibition was neither affected by the degree of the channel activation nor influenced by the types of divalent cations used for the channel activation. These results suggest that the NFA inhibition of ANO1 is likely mediated by altering the pore function but not through changing the channel gating. Our study provides a precise characterization of ANO1 and documents factors that can affect divalent cation activation and NFA inhibition of ANO1.

  14. Activation and Inhibition of TMEM16A Calcium-Activated Chloride Channels

    PubMed Central

    Ni, Yu-Li; Kuan, Ai-Seon; Chen, Tsung-Yu

    2014-01-01

    Calcium-activated chloride channels (CaCC) encoded by family members of transmembrane proteins of unknown function 16 (TMEM16) have recently been intensely studied for functional properties as well as their physiological roles as chloride channels in various tissues. One technical hurdle in studying these channels is the well-known channel rundown that frequently impairs the precision of electrophysiological measurements for the channels. Using experimental protocols that employ fast-solution exchange, we circumvented the problem of channel rundown by normalizing the Ca2+-induced current to the maximally-activated current obtained within a time period in which the channel rundown was negligible. We characterized the activation of the TMEM16A-encoded CaCC (also called ANO1) by Ca2+, Sr2+, and Ba2+, and discovered that Mg2+ competes with Ca2+ in binding to the divalent-cation binding site without activating the channel. We also studied the permeability of the ANO1 pore for various anions and found that the anion occupancy in the pore–as revealed by the permeability ratios of these anions–appeared to be inversely correlated with the apparent affinity of the ANO1 inhibition by niflumic acid (NFA). On the other hand, the NFA inhibition was neither affected by the degree of the channel activation nor influenced by the types of divalent cations used for the channel activation. These results suggest that the NFA inhibition of ANO1 is likely mediated by altering the pore function but not through changing the channel gating. Our study provides a precise characterization of ANO1 and documents factors that can affect divalent cation activation and NFA inhibition of ANO1. PMID:24489780

  15. Active and passive calcium transport systems in plant cells: Progress report, January 1986--June 1989

    SciTech Connect

    Sze, H.

    1989-01-01

    The objectives of this proposal are to identify and characterize active (energy-dependent) and passive calcium transport systems that work together to regulate calcium levels in the cytoplasm of plant cells. Several different energy-dependent Ca transport systems have been identified and characterized from oat root tissue (a monocot tissue) and carrot suspension cells (a dicot tissue). They are described in more detail below. I also have included in this progress report our continuing studies to understand the mode of action of the Helminthosporium maydis T toxin. This study was initially supported by a preceding DOE grant. The time needed to complete the study overlapped partly with the present grant period.

  16. Determination of the active center in calcium-nickel phosphate dehydrogenation catalyst

    SciTech Connect

    Attali, S.; Vigouroux, B.; Lenzi, M.; Pescia, J.

    1980-06-01

    Determination of the active center in calcium-nickel phosphate dehydrogenation catalyst, used industrially in the dehydrogenation of butenes to butadiene, showed that a stable trivalent nickel ion is involved. Apparently, electrons generated in the first (oxidation) step of the reaction are eliminated by reducing the trivalent to divalent nickel which is reoxidized by protons. The results were obtained by propanol dehydration-dehydrogenation on calcium-nickel phosphate (Ca/sub 8/Ni(PO/sub 4//sub )/6) calcined at 400/sup 0/-900/sup 0/C and by ESR spectroscopy.

  17. Adhesion of stainable, calcium-rich deposits on substrata with different surface free energies. An in vivo study in beagle dogs.

    PubMed

    Uyen, H M; van Dijk, L J; Busscher, H J

    1989-08-01

    Calculus plays an important role in chronic inflammatory periodontal disease and tooth loss. Patients can suffer from calculus formation despite good oral hygiene. The strength of adhesion between calculus and the enamel surface, though not determinant for the formation of calculus itself, determines whether calculus remains on the teeth during eating and toothbrushing. In this study, the amount and the strength of adhesion of calcium-rich deposits formed in vivo on different materials are related to substrate surface free energies (sfe). In 4 beagle dogs, fenestrated crowns were made on the upper fourth premolars. Smooth facings of glass (sfe 120 mJ.m-2), polished bovine enamel (sfe 85 mJ.m-2), polymethylmethacrylate (PMMA sfe 56 mJ.m-2) and polytetafluorethylene (PTFE sfe 20 mJ.m-2) were inserted in the crowns for 1, 3, 7, 14 or 28 days. The amount of deposit was evaluated both gravimetrically and planimetrically using Alizarin Red S for staining. Adhesion of calcium-rich deposits was evaluated planimetrically by studying their removal in a brushing machine. Dry weight increased linearly with time (approximately 0.18 mg.cm-2 per day) and was slightly less on PTFE than on the other materials. After 1 to 3 days, staining already revealed a 100% coverage by calcium-rich deposits. The number of strokes required to reduce the planimetric scores by 63% was extremely small on PTFE and PMMA and related with substrate surface free energies. This study shows that a possible way to reduce calculus formation in vivo is to decrease the surface free energy of the enamel using appropriate surfactants in, e.g., toothpastes.

  18. Oxalate Blockage of Calcium and Iron: A Student Learning Activity.

    ERIC Educational Resources Information Center

    Walker, Noojin

    1988-01-01

    Describes a student learning activity used to teach the meaning of percentage composition, mole concept, selective precipitation, and limiting factors. Presents two word problems and their solutions. (CW)

  19. Further characterization of ATP-initiated calcification by matrix vesicles isolated from rachitic rat cartilage. Membrane perturbation by detergents and deposition of calcium pyrophosphate by rachitic matrix vesicles.

    PubMed

    Hsu, H H; Camacho, N P; Anderson, H C

    1999-01-12

    Although membrane associated enzymes such as ATPase, alkaline phosphatase, and NTP pyrophosphohydrolase in matrix vesicles (MVs) may underlie the mechanisms of ATP-promoted calcification, prior to the current investigation, the role of the MV membrane in calcification had not been addressed. In this study, various perturbations were introduced to the MV membrane in in vitro calcification systems to determine ideal conditions for ATP-initiated calcification by MVs isolated from rachitic rat epiphyseal cartilage. Membrane integrity appears to be required, since the rupture of the vesicular membrane by vigorously mixing with 10% butanol abolished calcification. In contrast, a mild treatment of MVs with low concentrations (e.g., 0.01%, which is much below the critical concentration for micelle formation) of either neutral Triton X-100 or anionic deoxycholate stimulated calcification by >2-fold, without inducing obvious changes in vesicular appearance. Fourier transform infrared spectroscopic studies were done to identify the mineral phase formed in these experiments. For the first time, rachitic MVs were shown to induce the formation of a calcium pyrophosphate dihydrate-like phase after their exposure to calcifying medium with 1 mM ATP. The integration of spectral areas indicated that calcification was enhanced by Triton X-100. The detergent effect was reversible and appeared to be not mediated through activation of ATPase, alkaline phosphatase, or ATP pyrophosphohydrolase. In contrast to neutral Triton X-100 and anionic deoxycholate, cationic cetyltrimethylammonium bromide inhibited both ATPase activity (I50=10 microM) and ATP-initiated calcification. These observations suggest that membrane perturbations can affect calcification and that the presence of NTP-pyrophosphohydrolase in MVs may play a role in the deposition of CaPPi in rachitic cartilage.

  20. Correlations in distribution and concentration of calcium, copper and iron with zinc in isolated extracellular deposits associated with age-related macular degeneration

    USGS Publications Warehouse

    Flinn, Jane M; Kakalec, Peter; Tappero, Ryan; Jones, Blair F.; Lengyel, Imre

    2014-01-01

    Zinc (Zn) is abundantly enriched in sub-retinal pigment epithelial (RPE) deposits, the hallmarks of age-related macular degeneration (AMD), and is thought to play a role in the formation of these deposits. However, it is not known whether Zn is the only metal relevant for sub-RPE deposit formation. Because of their involvement in the pathogenesis of AMD, we determined the concentration and distribution of calcium (Ca), iron (Fe) and copper (Cu) and compared these with Zn in isolated and sectioned macular (MSD), equatorial (PHD) and far peripheral (FPD) sub-RPE deposits from an 86 year old donor eye with post mortem diagnosis of early AMD. The sections were mounted on Zn free microscopy slides and analyzed by microprobe synchrotron X-ray fluorescence (μSXRF). Metal concentrations were determined using spiked sectioned sheep brain matrix standards, prepared the same way as the samples. The heterogeneity of metal distributions was examined using pixel by pixel comparison. The orders of metal concentrations were Ca ⋙ Zn > Fe in all three types of deposits but Cu levels were not distinguishable from background values. Zinc and Ca were consistently present in all deposits but reached highest concentration in MSD. Iron was present in some but not all deposits and was especially enriched in FPD. Correlation analysis indicated considerable variation in metal distribution within and between sub-RPE deposits. The results suggest that Zn and Ca are the most likely contributors to deposit formation especially in MSD, the characteristic risk factor for the development of AMD in the human eye.

  1. Activation of Oral Trigeminal Neurons by Fatty Acids is Dependent upon Intracellular Calcium

    PubMed Central

    Yu, Tian; Shah, Bhavik P.; Hansen, Dane R.; Park-York, MieJung; Gilbertson, Timothy A.

    2012-01-01

    The chemoreception of dietary fat in the oral cavity has largely been attributed to activation of the somatosensory system that conveys the textural properties of fat. However, the ability of fatty acids, which are believed to represent the proximate stimulus for fat taste, to stimulate rat trigeminal neurons has remained unexplored. Here, we found that several free fatty acids are capable of activating trigeminal neurons with different kinetics. Further, a polyunsaturated fatty acid, linoleic acid (LA), activates trigeminal neurons by increasing intracellular calcium concentration and generating depolarizing receptor potentials. Ion substitution and pharmacological approaches reveal that intracellular calcium store depletion is crucial for LA-induced signaling in a subset of trigeminal neurons. Using pseudorabies virus (PrV) as a live cell tracer, we identified a subset of lingual nerve-innervated trigeminal neurons that respond to different subsets of fatty acids. Quantitative real-time PCR of several transient receptor potential (TRP) channel markers in individual neurons validated that PrV labeled a subset but not the entire population of lingual-innervated trigeminal neurons. We further confirmed that the LA-induced intracellular calcium rise is exclusively coming from the release of calcium stores from the endoplasmic reticulum in this subset of lingual nerve-innervated trigeminal neurons. PMID:22644615

  2. Where is the origin of the activator calcium in cardiac ventricular contraction?

    PubMed

    Reiter, M; Vierling, W; Seibel, K

    1984-01-01

    Under normal experimental conditions, the force of rested-state contractions (i.e., contractions after a rest period of 15 min or longer) of mammalian ventricular myocardium is insignificant. In Mg2+-free solution, in low sodium solution or in the presence of a cardioactive steroid, a strong "early" rested-state contraction develops without delay after stimulation, indicating the accumulation during rest of intracellularly stored activator calcium. By contrast, catecholamines cause a "late" rested-state contraction with a characteristic latent period of about 100 ms between stimulation and onset of contraction. Inhibition of the slow inward current by nifedipine has no influence on the contraction velocity of the "early" rested-state contraction, indicating that Ca2+ of the slow inward current is not involved in the calcium release mechanism of prefilled stores during excitation-contraction coupling. Nifedipine suppresses the "late" rested-state contraction in the presence of noradrenaline. In view of the constancy of the latent period, it is proposed that the activator calcium for the "late" rested-state contraction enters the cell with the slow inward current, is sequestered at first by uptake sites of the sarcoplasmic reticulum and subsequently released from its release sites as long as the cell is depolarized. The model of the different origin of activator calcium is discussed in its implication for high-frequency contractions.

  3. 3-Bromopyruvate inhibits calcium uptake by sarcoplasmic reticulum vesicles but not SERCA ATP hydrolysis activity.

    PubMed

    Jardim-Messeder, Douglas; Camacho-Pereira, Juliana; Galina, Antonio

    2012-05-01

    3-Bromopyruvate (3BrPA) is an antitumor agent that alkylates the thiol groups of enzymes and has been proposed as a treatment for neoplasias because of its specific reactivity with metabolic energy transducing enzymes in tumor cells. In this study, we show that the sarco/endoplasmic reticulum calcium (Ca(2+)) ATPase (SERCA) type 1 is one of the target enzymes of 3BrPA activity. Sarco/endoplasmic reticulum vesicles (SRV) were incubated in the presence of 1mM 3BrPA, which was unable to inhibit the ATPase activity of SERCA. However, Ca(2+)-uptake activity was significantly inhibited by 80% with 150 μM 3BrPA. These results indicate that 3BrPA has the ability to uncouple the ATP hydrolysis from the calcium transport activities. In addition, we observed that the inclusion of 2mM reduced glutathione (GSH) in the reaction medium with different 3BrPA concentrations promoted an increase in 40% in ATPase activity and protects the inhibition promoted by 3BrPA in calcium uptake activity. This derivatization is accompanied by a decrease of reduced cysteine (Cys), suggesting that GSH and 3BrPA increases SERCA activity and transport by pyruvylation and/or S-glutathiolation mediated by GSH at a critical Cys residues of the SERCA.

  4. Effects of Calcium Spikes in the Layer 5 Pyramidal Neuron on Coincidence Detection and Activity Propagation

    PubMed Central

    Chua, Yansong; Morrison, Abigail

    2016-01-01

    The role of dendritic spiking mechanisms in neural processing is so far poorly understood. To investigate the role of calcium spikes in the functional properties of the single neuron and recurrent networks, we investigated a three compartment neuron model of the layer 5 pyramidal neuron with calcium dynamics in the distal compartment. By performing single neuron simulations with noisy synaptic input and occasional large coincident input at either just the distal compartment or at both somatic and distal compartments, we show that the presence of calcium spikes confers a substantial advantage for coincidence detection in the former case and a lesser advantage in the latter. We further show that the experimentally observed critical frequency phenomenon, in which action potentials triggered by stimuli near the soma above a certain frequency trigger a calcium spike at distal dendrites, leading to further somatic depolarization, is not exhibited by a neuron receiving realistically noisy synaptic input, and so is unlikely to be a necessary component of coincidence detection. We next investigate the effect of calcium spikes in propagation of spiking activities in a feed-forward network (FFN) embedded in a balanced recurrent network. The excitatory neurons in the network are again connected to either just the distal, or both somatic and distal compartments. With purely distal connectivity, activity propagation is stable and distinguishable for a large range of recurrent synaptic strengths if the feed-forward connections are sufficiently strong, but propagation does not occur in the absence of calcium spikes. When connections are made to both the somatic and the distal compartments, activity propagation is achieved for neurons with active calcium dynamics at a much smaller number of neurons per pool, compared to a network of passive neurons, but quickly becomes unstable as the strength of recurrent synapses increases. Activity propagation at higher scaling factors can be

  5. Effects of Calcium Spikes in the Layer 5 Pyramidal Neuron on Coincidence Detection and Activity Propagation.

    PubMed

    Chua, Yansong; Morrison, Abigail

    2016-01-01

    The role of dendritic spiking mechanisms in neural processing is so far poorly understood. To investigate the role of calcium spikes in the functional properties of the single neuron and recurrent networks, we investigated a three compartment neuron model of the layer 5 pyramidal neuron with calcium dynamics in the distal compartment. By performing single neuron simulations with noisy synaptic input and occasional large coincident input at either just the distal compartment or at both somatic and distal compartments, we show that the presence of calcium spikes confers a substantial advantage for coincidence detection in the former case and a lesser advantage in the latter. We further show that the experimentally observed critical frequency phenomenon, in which action potentials triggered by stimuli near the soma above a certain frequency trigger a calcium spike at distal dendrites, leading to further somatic depolarization, is not exhibited by a neuron receiving realistically noisy synaptic input, and so is unlikely to be a necessary component of coincidence detection. We next investigate the effect of calcium spikes in propagation of spiking activities in a feed-forward network (FFN) embedded in a balanced recurrent network. The excitatory neurons in the network are again connected to either just the distal, or both somatic and distal compartments. With purely distal connectivity, activity propagation is stable and distinguishable for a large range of recurrent synaptic strengths if the feed-forward connections are sufficiently strong, but propagation does not occur in the absence of calcium spikes. When connections are made to both the somatic and the distal compartments, activity propagation is achieved for neurons with active calcium dynamics at a much smaller number of neurons per pool, compared to a network of passive neurons, but quickly becomes unstable as the strength of recurrent synapses increases. Activity propagation at higher scaling factors can be

  6. The effects of therapeutic exercise using PNF on the size of calcium deposits, pain self-awareness, and shoulder joint function in a calcific tendinitis patient: a case study

    PubMed Central

    Oh, Dong-Gun; Yoo, Kyung-Tae

    2017-01-01

    [Purpose] The purpose of this case study was to identify the effects of independent and intensive therapeutic exercise using Proprioceptive neuromuscular facilitation on the size of calcium deposits, pain self-awareness, and shoulder joint function in a patient with calcific tendonitis. [Subject and Methods] The subject was a 42-year-old female patient with calcific tendonitis and acute pain who had difficulty with active movement and problems with general function. The independent and intensive Proprioceptive neuromuscular facilitation exercise was applied for 40 min twice a day five times a week for two weeks for a total of 20 times. An X-ray, the visual analog scale, a simple shoulder test, the Constant-Murley Scale, and passive range of motion was used to evaluate the patient’s change. [Results] The size of the calcium deposit, the visual analog scale score, and the simple shoulder test score decreased. The Constant-Murley Scale score and the passive range of motion were increased. [Conclusion] The results of this study suggested that intensive and independent therapeutic short-term exercise without any other exercise reduced pain and produced positive effects in shoulder function in a patient with the calcific tendonitis, which could confirm the importance of therapeutic exercise in the treatment of calcific tendonitis. PMID:28210065

  7. Calcium-dependent activation of mitochondrial metabolism in mammalian cells

    PubMed Central

    Gaspers, Lawrence D.; Thomas, Andrew P.

    2008-01-01

    Endogenous fluorophores provide a simple, but elegant means to investigate the relationship between agonist-evoked Ca2+ signals and the activation of mitochondrial metabolism. In this article, we discuss the methods and strategies to measure cellular pyridine nucleotide and flavoprotein fluorescence alone or in combination with Ca2+-sensitive indicators. These methods were developed using primary cultured hepatocytes and neurons, which contain relatively high levels of endogenous fluorophores and robust metabolic responses. Nevertheless, these methods are amendable to a wide variety of primary cell types and cell lines that maintain active mitochondrial metabolism. PMID:18854213

  8. Calcium-activated chloride current expression in axotomized sensory neurons: what for?

    PubMed Central

    Boudes, Mathieu; Scamps, Frédérique

    2012-01-01

    Calcium-activated chloride currents (CaCCs) are activated by an increase in intracellular calcium concentration. Peripheral nerve injury induces the expression of CaCCs in a subset of adult sensory neurons in primary culture including mechano- and proprioceptors, though not nociceptors. Functional screenings of potential candidate genes established that Best1 is a molecular determinant for CaCC expression among axotomized sensory neurons, while Tmem16a is acutely activated by inflammatory mediators in nociceptors. In nociceptors, such CaCCs are preferentially activated under receptor-induced calcium mobilization contributing to cell excitability and pain. In axotomized mechano- and proprioceptors, CaCC activation does not promote electrical activity and prevents firing, a finding consistent with electrical silencing for growth competence of adult sensory neurons. In favor of a role in the process of neurite growth, CaCC expression is temporally correlated to neurons displaying a regenerative mode of growth. This perspective focuses on the molecular identity and role of CaCC in axotomized sensory neurons and the future directions to decipher the cellular mechanisms regulating CaCC during neurite (re)growth. PMID:22461766

  9. Calcium-activated chloride channels anoctamin 1 and 2 promote murine uterine smooth muscle contractility

    PubMed Central

    Bernstein, Kyra; Vink, Joy Y; Fu, Xiao Wen; Wakita, Hiromi; Danielsson, Jennifer; Wapner, Ronald; Gallos, George

    2014-01-01

    Objective To determine the presence of calcium activated chloride channels anoctamin 1 and 2 in human and murine uterine smooth muscle and evaluate the physiologic role for these ion channels in murine myometrial contractility. Study Design We performed reverse transcription polymerase chain reaction to determine if anoctamin 1 and 2 are expressed in human and murine uterine tissue to validate the study of this protein in mouse models. Immunohistochemical staining of anoctamin 1 and 2 was then performed to determine protein expression in murine myometrial tissue. The function of anoctamin 1 and 2 in murine uterine tissue was evaluated using electrophysiological studies, organ bath, and calcium flux experiments. Results Anoctamin 1 and 2 are expressed in human and murine USM cells. Functional studies show that selective antagonism of these channels promotes relaxation of spontaneous murine uterine smooth muscle contractions. Blockade of anoctamin 1 and 2 inhibits both agonist-induced and spontaneous transient inward currents and abolishes G-protein coupled receptor (oxytocin) mediated elevations in intracellular calcium. Conclusion The calcium activated chloride channels ANO 1 and 2 are present in human and murine myometrial tissue and may provide novel potential therapeutic targets to achieve effective tocolysis. PMID:24928056

  10. Oxidative Regulation of Large Conductance Calcium-Activated Potassium Channels

    PubMed Central

    Tang, Xiang D.; Daggett, Heather; Hanner, Markus; Garcia, Maria L.; McManus, Owen B.; Brot, Nathan; Weissbach, Herbert; Heinemann, Stefan H.; Hoshi, Toshinori

    2001-01-01

    Reactive oxygen/nitrogen species are readily generated in vivo, playing roles in many physiological and pathological conditions, such as Alzheimer's disease and Parkinson's disease, by oxidatively modifying various proteins. Previous studies indicate that large conductance Ca2+-activated K+ channels (BKCa or Slo) are subject to redox regulation. However, conflicting results exist whether oxidation increases or decreases the channel activity. We used chloramine-T, which preferentially oxidizes methionine, to examine the functional consequences of methionine oxidation in the cloned human Slo (hSlo) channel expressed in mammalian cells. In the virtual absence of Ca2+, the oxidant shifted the steady-state macroscopic conductance to a more negative direction and slowed deactivation. The results obtained suggest that oxidation enhances specific voltage-dependent opening transitions and slows the rate-limiting closing transition. Enhancement of the hSlo activity was partially reversed by the enzyme peptide methionine sulfoxide reductase, suggesting that the upregulation is mediated by methionine oxidation. In contrast, hydrogen peroxide and cysteine-specific reagents, DTNB, MTSEA, and PCMB, decreased the channel activity. Chloramine-T was much less effective when concurrently applied with the K+ channel blocker TEA, which is consistent with the possibility that the target methionine lies within the channel pore. Regulation of the Slo channel by methionine oxidation may represent an important link between cellular electrical excitability and metabolism. PMID:11222629

  11. N-cadherin modulates voltage activated calcium influx via RhoA, p120-catenin, and myosinactin interaction

    PubMed Central

    Marrs, Glen S.; Theisen, Christopher S.; Brusés, Juan L.

    2010-01-01

    N-cadherin is a transmembrane adhesion receptor that contributes to neuronal development and synapse formation through homophilic interactions that provide structural-adhesive support to contacts between cell membranes. In addition, N-cadherin homotypic binding may initiate cell signaling that regulates neuronal physiology. In this study, we investigated signaling capabilities of N-cadherin that control voltage activated calcium influx. Using whole-cell voltage clamp recording of isolated inward calcium currents in freshly isolated chick ciliary ganglion neurons we show that the juxtamembrane region of N-cadherin cytoplasmic domain regulates high-threshold voltage activated calcium currents by interacting with p120-catenin and activating RhoA. This regulatory mechanism requires myosin interaction with actin. Furthermore, N-cadherin homophilic binding enhanced voltage activated calcium current amplitude in dissociated neurons that have already developed mature synaptic contacts in vivo. The increase in calcium current amplitude was not affected by brefeldin A suggesting that the effect is caused via direct channel modulation and not by increasing channel expression. In contrast, homotypic N-cadherin interaction failed to regulate calcium influx in freshly isolated immature neurons. However, RhoA inhibitors enhanced calcium current amplitude in these immature neurons, suggesting that the inhibitory effect of RhoA on calcium entry is regulated during neuronal development and synapse maturation. These results indicate that N-cadherin modulates voltage activated calcium entry by a mechanism that involves RhoA activity and its downstream effects on the cytoskeleton, and suggest that N-cadherin provides support for synaptic maturation and sustained synaptic activity by facilitating voltage activated calcium influx. PMID:19162191

  12. Stimulus-response coupling in human platelets. Changes evoked by platelet-activating factor in cytoplasmic free calcium monitored with the fluorescent calcium indicator quin2.

    PubMed Central

    Hallam, T J; Sanchez, A; Rink, T J

    1984-01-01

    The role of changes in cytoplasmic free calcium, [Ca2+]i, in the responses to platelet-activating factor (PAF) was studied in human platelets loaded with the fluorescent calcium indicator, quin2. In the presence of 1 mM external calcium, PAF raised [Ca2+]i 8-10-fold in a few seconds to peak near 1 microM. [Ca2+]i then declined over several minutes towards the basal level. In the absence of external calcium there was a much smaller increase in [Ca2+]i of similar pattern. These findings suggest that PAF increases [Ca2+]i partly by discharge of internal Ca2+, but mainly by stimulated influx. Blockade of cyclo-oxygenase with aspirin only slightly reduced the [Ca2+]i changes, indicating that thromboxane A2 is not a major mediator of the calcium movements. In control conditions PAF could stimulate shape-change, aggregation and secretion. Aggregation and secretion were roughly halved by blockade of cyclo-oxygenase. Shape-change and secretion still occurred under conditions where the [Ca2+]i rise was small or suppressed, indicating a role for intracellular activators other than Ca2+. The possible involvement of products of phosphoinositide breakdown is discussed. PMID:6426464

  13. Synaptic vesicle exocytosis and increased cytosolic calcium are both necessary but not sufficient for activity-dependent bulk endocytosis.

    PubMed

    Morton, Andrew; Marland, Jamie R K; Cousin, Michael A

    2015-08-01

    Activity-dependent bulk endocytosis (ADBE) is the dominant synaptic vesicle (SV) endocytosis mode in central nerve terminals during intense neuronal activity. By definition this mode is triggered by neuronal activity; however, key questions regarding its mechanism of activation remain unaddressed. To determine the basic requirements for ADBE triggering in central nerve terminals, we decoupled SV fusion events from activity-dependent calcium influx using either clostridial neurotoxins or buffering of intracellular calcium. ADBE was monitored both optically and morphologically by observing uptake of the fluid phase markers tetramethylrhodamine-dextran and horse radish peroxidase respectively. Ablation of SV fusion with tetanus toxin resulted in the arrest of ADBE, but had no effect on other calcium-dependent events such as activity-dependent dynamin I dephosphorylation, indicating that SV exocytosis is necessary for triggering. Furthermore, the calcium chelator EGTA abolished ADBE while leaving SV exocytosis intact, demonstrating that ADBE is triggered by intracellular free calcium increases outside the active zone. Activity-dependent dynamin I dephosphorylation was also arrested in EGTA-treated neurons, consistent with its proposed role in triggering ADBE. Thus, SV fusion and increased cytoplasmic free calcium are both necessary but not sufficient individually to trigger ADBE. Activity-dependent bulk endocytosis (ADBE) is the dominant synaptic vesicle (SV) endocytosis mode in central nerve terminals during intense neuronal activity. To determine the minimal requirements for ADBE triggering, we decoupled SV fusion events from activity-dependent calcium influx using either clostridial neurotoxins or buffering of intracellular calcium. We found that SV fusion and increased cytoplasmic free calcium are both necessary but not sufficient to trigger ADBE.

  14. Tissue plasminogen activator inhibits NMDA-receptor-mediated increases in calcium levels in cultured hippocampal neurons

    PubMed Central

    Robinson, Samuel D.; Lee, Tet Woo; Christie, David L.; Birch, Nigel P.

    2015-01-01

    NMDA receptors (NMDARs) play a critical role in neurotransmission, acting as essential mediators of many forms of synaptic plasticity, and also modulating aspects of development, synaptic transmission and cell death. NMDAR-induced responses are dependent on a range of factors including subunit composition and receptor location. Tissue-type plasminogen activator (tPA) is a serine protease that has been reported to interact with NMDARs and modulate NMDAR activity. In this study we report that tPA inhibits NMDAR-mediated changes in intracellular calcium levels in cultures of primary hippocampal neurons stimulated by low (5 μM) but not high (50 μM) concentrations of NMDA. tPA also inhibited changes in calcium levels stimulated by presynaptic release of glutamate following treatment with bicucculine/4-aminopyridine (4-AP). Inhibition was dependent on the proteolytic activity of tPA but was unaffected by α2-antiplasmin, an inhibitor of the tPA substrate plasmin, and receptor-associated protein (RAP), a pan-ligand blocker of the low-density lipoprotein receptor, two proteins previously reported to modulate NMDAR activity. These findings suggest that tPA can modulate changes in intracellular calcium levels in a subset of NMDARs expressed in cultured embryonic hippocampal neurons through a mechanism that involves the proteolytic activity of tPA and synaptic NMDARs. PMID:26500501

  15. Calmodulin-dependent activation and inactivation of anoctamin calcium-gated chloride channels.

    PubMed

    Vocke, Kerstin; Dauner, Kristin; Hahn, Anne; Ulbrich, Anne; Broecker, Jana; Keller, Sandro; Frings, Stephan; Möhrlen, Frank

    2013-10-01

    Calcium-dependent chloride channels serve critical functions in diverse biological systems. Driven by cellular calcium signals, the channels codetermine excitatory processes and promote solute transport. The anoctamin (ANO) family of membrane proteins encodes three calcium-activated chloride channels, named ANO 1 (also TMEM16A), ANO 2 (also TMEM16B), and ANO 6 (also TMEM16F). Here we examined how ANO 1 and ANO 2 interact with Ca(2+)/calmodulin using nonstationary current analysis during channel activation. We identified a putative calmodulin-binding domain in the N-terminal region of the channel proteins that is involved in channel activation. Binding studies with peptides indicated that this domain, a regulatory calmodulin-binding motif (RCBM), provides two distinct modes of interaction with Ca(2+)/calmodulin, one at submicromolar Ca(2+) concentrations and one in the micromolar Ca(2+) range. Functional, structural, and pharmacological data support the concept that calmodulin serves as a calcium sensor that is stably associated with the RCBM domain and regulates the activation of ANO 1 and ANO 2 channels. Moreover, the predominant splice variant of ANO 2 in the brain exhibits Ca(2+)/calmodulin-dependent inactivation, a loss of channel activity within 30 s. This property may curtail ANO 2 activity during persistent Ca(2+) signals in neurons. Mutagenesis data indicated that the RCBM domain is also involved in ANO 2 inactivation, and that inactivation is suppressed in the retinal ANO 2 splice variant. These results advance the understanding of Ca(2+) regulation in anoctamin Cl(-) channels and its significance for the physiological function that anoctamin channels subserve in neurons and other cell types.

  16. Calmodulin-dependent activation and inactivation of anoctamin calcium-gated chloride channels

    PubMed Central

    Vocke, Kerstin; Dauner, Kristin; Hahn, Anne; Ulbrich, Anne; Broecker, Jana; Keller, Sandro; Frings, Stephan

    2013-01-01

    Calcium-dependent chloride channels serve critical functions in diverse biological systems. Driven by cellular calcium signals, the channels codetermine excitatory processes and promote solute transport. The anoctamin (ANO) family of membrane proteins encodes three calcium-activated chloride channels, named ANO 1 (also TMEM16A), ANO 2 (also TMEM16B), and ANO 6 (also TMEM16F). Here we examined how ANO 1 and ANO 2 interact with Ca2+/calmodulin using nonstationary current analysis during channel activation. We identified a putative calmodulin-binding domain in the N-terminal region of the channel proteins that is involved in channel activation. Binding studies with peptides indicated that this domain, a regulatory calmodulin-binding motif (RCBM), provides two distinct modes of interaction with Ca2+/calmodulin, one at submicromolar Ca2+ concentrations and one in the micromolar Ca2+ range. Functional, structural, and pharmacological data support the concept that calmodulin serves as a calcium sensor that is stably associated with the RCBM domain and regulates the activation of ANO 1 and ANO 2 channels. Moreover, the predominant splice variant of ANO 2 in the brain exhibits Ca2+/calmodulin-dependent inactivation, a loss of channel activity within 30 s. This property may curtail ANO 2 activity during persistent Ca2+ signals in neurons. Mutagenesis data indicated that the RCBM domain is also involved in ANO 2 inactivation, and that inactivation is suppressed in the retinal ANO 2 splice variant. These results advance the understanding of Ca2+ regulation in anoctamin Cl− channels and its significance for the physiological function that anoctamin channels subserve in neurons and other cell types. PMID:24081981

  17. Amphetamine activates calcium channels through dopamine transporter-mediated depolarization.

    PubMed

    Cameron, Krasnodara N; Solis, Ernesto; Ruchala, Iwona; De Felice, Louis J; Eltit, Jose M

    2015-11-01

    Amphetamine (AMPH) and its more potent enantiomer S(+)AMPH are psychostimulants used therapeutically to treat attention deficit hyperactivity disorder and have significant abuse liability. AMPH is a dopamine transporter (DAT) substrate that inhibits dopamine (DA) uptake and is implicated in DA release. Furthermore, AMPH activates ionic currents through DAT that modify cell excitability presumably by modulating voltage-gated channel activity. Indeed, several studies suggest that monoamine transporter-induced depolarization opens voltage-gated Ca(2+) channels (CaV), which would constitute an additional AMPH mechanism of action. In this study we co-express human DAT (hDAT) with Ca(2+) channels that have decreasing sensitivity to membrane depolarization (CaV1.3, CaV1.2 or CaV2.2). Although S(+)AMPH is more potent than DA in transport-competition assays and inward-current generation, at saturating concentrations both substrates indirectly activate voltage-gated L-type Ca(2+) channels (CaV1.3 and CaV1.2) but not the N-type Ca(2+) channel (CaV2.2). Furthermore, the potency to achieve hDAT-CaV electrical coupling is dominated by the substrate affinity on hDAT, with negligible influence of L-type channel voltage sensitivity. In contrast, the maximal coupling-strength (defined as Ca(2+) signal change per unit hDAT current) is influenced by CaV voltage sensitivity, which is greater in CaV1.3- than in CaV1.2-expressing cells. Moreover, relative to DA, S(+)AMPH showed greater coupling-strength at concentrations that induced relatively small hDAT-mediated currents. Therefore S(+)AMPH is not only more potent than DA at inducing hDAT-mediated L-type Ca(2+) channel currents but is a better depolarizing agent since it produces tighter electrical coupling between hDAT-mediated depolarization and L-type Ca(2+) channel activation.

  18. Bone Mineral Density Changes after Physical Training and Calcium Intake in Students with Attention Deficit and Hyper Activity Disorders

    ERIC Educational Resources Information Center

    Arab ameri, Elahe; Dehkhoda, Mohammad Reza; Hemayattalab, Rasool

    2012-01-01

    In this study we investigate the effects of weight bearing exercise and calcium intake on bone mineral density (BMD) of students with attention deficit and hyper activity (ADHD) disorder. For this reason 54 male students with ADHD (age 8-12 years old) were assigned to four groups with no differences in age, BMD, calcium intake, and physical…

  19. Boron Accelerates Cultured Osteoblastic Cell Activity through Calcium Flux.

    PubMed

    Capati, Mark Luigi Fabian; Nakazono, Ayako; Igawa, Kazunari; Ookubo, Kensuke; Yamamoto, Yuya; Yanagiguchi, Kajirou; Kubo, Shisei; Yamada, Shizuka; Hayashi, Yoshihiko

    2016-12-01

    A low concentration of boron (B) accelerates the proliferation and differentiation of mammalian osteoblasts. The aim of this study was to investigate the effects of 0.1 mM of B on the membrane function of osteoblastic cells in vitro. Genes involved in cell activity were investigated using gene expression microarray analyses. The Ca(2+) influx and efflux were evaluated to demonstrate the activation of L-type Ca(2+) channel for the Ca(2+) influx, and that of Na(+)/K(+)-ATPase for the Ca(2+) efflux. A real-time PCR analysis revealed that the messenger RNA (mRNA) expression of four mineralization-related genes was clearly increased after 3 days of culture with a B-supplemented culture medium. Using microarray analyses, five genes involved in cell proliferation and differentiation were upregulated compared to the control group. Regarding the Ca(2+) influx, in the nifedipine-pretreated group, the relative fluorescence intensity for 1 min after adding B solution did not increase compared with that for 1 min before addition. In the control group, the relative fluorescence intensity was significantly increased compared with the experimental group (P < 0.05). Regarding the Ca(2+) efflux, in the experimental group cultured in 0.1 mM of B-supplemented medium, the relative fluorescence intensity for 10 min after ouabain treatment revealed a significantly lower slope value compared with the control group (P < 0.01). This is the first study to demonstrate the acceleration of Ca(2+) flux by B supplementation in osteoblastic cells. Cell membrane stability is related to the mechanism by which a very low concentration of B promotes the proliferation and differentiation of mammalian osteoblastic cells in vitro.

  20. Cytosolic calcium coordinates mitochondrial energy metabolism with presynaptic activity.

    PubMed

    Chouhan, Amit K; Ivannikov, Maxim V; Lu, Zhongmin; Sugimori, Mutsuyuki; Llinas, Rodolfo R; Macleod, Gregory T

    2012-01-25

    Most neurons fire in bursts, imposing episodic energy demands, but how these demands are coordinated with oxidative phosphorylation is still unknown. Here, using fluorescence imaging techniques on presynaptic termini of Drosophila motor neurons (MNs), we show that mitochondrial matrix pH (pHm), inner membrane potential (Δψm), and NAD(P)H levels ([NAD(P)H]m) increase within seconds of nerve stimulation. The elevations of pHm, Δψm, and [NAD(P)H]m indicate an increased capacity for ATP production. Elevations in pHm were blocked by manipulations that blocked mitochondrial Ca2+ uptake, including replacement of extracellular Ca2+ with Sr2+ and application of either tetraphenylphosphonium chloride or KB-R7943, indicating that it is Ca2+ that stimulates presynaptic mitochondrial energy metabolism. To place this phenomenon within the context of endogenous neuronal activity, the firing rates of a number of individually identified MNs were determined during fictive locomotion. Surprisingly, although endogenous firing rates are significantly different, there was little difference in presynaptic cytosolic Ca2+ levels ([Ca2+]c) between MNs when each fires at its endogenous rate. The average [Ca2+]c level (329±11 nM) was slightly above the average Ca2+ affinity of the mitochondria (281±13 nM). In summary, we show that when MNs fire at endogenous rates, [Ca2+]c is driven into a range where mitochondria rapidly acquire Ca2+. As we also show that Ca2+ stimulates presynaptic mitochondrial energy metabolism, we conclude that [Ca2+]c levels play an integral role in coordinating mitochondrial energy metabolism with presynaptic activity in Drosophila MNs.

  1. Cytosolic Calcium Coordinates Mitochondrial Energy Metabolism with Presynaptic Activity

    PubMed Central

    Chouhan, Amit K.; Ivannikov, Maxim V.; Lu, Zhongmin; Sugimori, Mutsuyuki; Llinas, Rodolfo R.; Macleod, Gregory T.

    2012-01-01

    Most neurons fire in bursts, imposing episodic energy demands, but how these demands are coordinated with oxidative phosphorylation is still unknown. Here, using fluorescence imaging techniques on presynaptic termini of Drosophila motor neurons (MNs), we show that mitochondrial matrix pH (pHm), inner membrane potential (Δψm), and NAD(P)H levels ([NAD(P)H]m) increase within seconds of nerve stimulation. The elevations of pHm, Δψm, and [NAD(P)H]m indicate an increased capacity for ATP production. Elevations in pHm were blocked by manipulations which blocked mitochondrial Ca2+ uptake, including replacement of extracellular Ca2+ with Sr2+, and application of either tetraphenylphosphonium chloride or KB-R7943, indicating that it is Ca2+ that stimulates presynaptic mitochondrial energy metabolism. To place this phenomenon within the context of endogenous neuronal activity, the firing rates of a number of individually identified MNs were determined during fictive locomotion. Surprisingly, although endogenous firing rates are significantly different, there was little difference in presynaptic cytosolic Ca2+ levels ([Ca2+]c) between MNs when each fires at its endogenous rate. The average [Ca2+]c level (329±11nM) was slightly above the average Ca2+ affinity of the mitochondria (281±13nM). In summary, we show that when MNs fire at endogenous rates [Ca2+]c is driven into a range where mitochondria rapidly acquire Ca2+. As we also show that Ca2+ stimulates presynaptic mitochondrial energy metabolism, we conclude that [Ca2+]c levels play an integral role in coordinating mitochondrial energy metabolism with presynaptic activity in Drosophila MNs. PMID:22279208

  2. Leptin regulated calcium channels of neuropeptide Y and proopiomelanocortin neurons by activation of different signal pathways.

    PubMed

    Wang, J-H; Wang, F; Yang, M-J; Yu, D-F; Wu, W-N; Liu, J; Ma, L-Q; Cai, F; Chen, J-G

    2008-09-22

    The fat-derived hormone leptin regulates food intake and body weight in part by modulating the activity of neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons in the hypothalamic arcuate nucleus (ARC). To investigate the electrophysiological activity of these neurons and their responses to leptin, we recorded whole-cell calcium currents on NPY and POMC neurons in the ARC of rats, which we identified by morphologic features and immunocytochemical identification at the end of recording. Leptin decreased the peak amplitude of high voltage-activated calcium currents (I(HVA)) in the isolated neurons from ARC, which were subsequently shown to be immunoreactive for NPY. The inhibition was prevented by pretreatment with inhibitors of Janus kinase 2 (JAK2) and mitogen-activated protein kinases (MAPK). In contrast, leptin increased the amplitude of I(HVA) in POMC-containing neurons. The stimulations of I(HVA) were inhibited by blockers of JAK2 and phosphatidylino 3-kinase (PI3-k). Both of these effects were counteracted by the L-type calcium channel antagonist nifedipine, suggesting that L-type calcium channels were involved in the regulation induced by leptin. These data indicated that leptin exerted opposite effects on these two classes of neurons. Leptin directly inhibited I(HVA) in NPY neurons via leptin receptor (LEPR) -JAK2-MAPK pathways, whereas evoked I(HVA) in POMC neurons by LEPR-JAK2-PI3-k pathways. These neural pathways and intracellular signaling mechanisms may play key roles in regulating NPY and POMC neuron activity, anorectic action of leptin and, thereby, feeding.

  3. Influences of Ag-NPs doping chitosan/calcium silicate nanocomposites for optical and antibacterial activity.

    PubMed

    El-Nahrawy, Amany M; Ali, Ahmed I; Abou Hammad, Ali B; Youssef, Ahmed M

    2016-12-01

    Chitosan (CS)/calcium silicate nanocomposites pure and doped with Ag ions (1, 2mol%) were prepared via sol-gel method. The prepared CS/calcium silicate nanocomposites were investigated through X-ray diffraction (XRD), Fourier transforms infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM). The XRD results, indicating that after increasing the Ag ions in the CS/calcium silicate nanocomposite the crystallinity degree increased regularly in the prepared nanocomposites accompanying to the continuously rearrangement in the internal structure of nanocomposite under the effect of inorganic nanoparticles. Correspondingly, the optical properties of the prepared nanocomposites films were measured using UV/vis spectroscopy. The reflectance increased while the energy band gap decreased from 3.96eV to 2.43eV with Ag-ions concentration. More over the transition type changed from direct into indirect by adding Ag-ions, indicate that new band between valence and conduction band were formed. In addition, the optical parameters showed an increase in refractive indices and decrease in the surface and volume energies losses with increasing Ag-ions. Correspondingly, the prepared nanocomposites exhibited good antibacterial activity against gram positive (Staphylococcus aureus), gram negative (Pseudomonas aeruginosa) bacteria and fungi (Candidia albicans). The results suggested that the prepared CS/calcium silicate nanocomposites can be a promised candidate for optical sensors applications and smart packaging materials.

  4. Synthesis and evaluation of 1,4-dihydropyridine derivatives with calcium channel blocking activity.

    PubMed

    Bladen, Chris; Gündüz, Miyase Gözde; Şimşek, Rahime; Şafak, Cihat; Zamponi, Gerald W

    2014-07-01

    1,4-Dihydropyridines (DHPs) are an important class of L-type calcium channel blockers that are used to treat conditions such as hypertension and angina. Their primary target in the cardiovascular system is the Cav1.2 L-type calcium channel isoform, however, a number of DHPs also block low-voltage-activated T-type calcium channels. Here, we describe the synthesis of a series of novel DHP derivatives that have a condensed 1,4-DHP ring system (hexahydroquinoline) and report on their abilities to block both L- and T-type calcium channels. Within this series of compounds, modification of a key ester moiety not only regulates the blocking affinity for both L- and T-type channels, but also allows for the development of DHPs with 30-fold selectivity for T-type channels over the L-type. Our data suggest that a condensed dihydropyridine-based scaffold may serve as a pharmacophore for a new class of T-type selective inhibitors.

  5. Role of low-voltage-activated calcium current and extracellular calcium in controlling the firing pattern of developing CA1 pyramidal neurons.

    PubMed

    Sánchez-Aguilera, Alberto; Sánchez-Alonso, José Luis; Vicente-Torres, María Ángeles; Colino, Asunción

    2017-03-06

    The firing pattern of individual neurons is an important element for information processing and storing. During the first weeks of development, there is a transitional period during which CA1 pyramidal neurons display burst-spiking behavior in contrast to the adult regular-firing pattern. Spike after-depolarizations (ADPs) constitute a major factor underlying burst-spiking behavior. Using current-clamp recordings, we studied ADP waveforms and firing patterns in CA1 pyramidal neurons of Wistar rats from 9 to 19 postnatal days (P9-19). The percentage of burst-spiking neurons increased up to P16, in correlation with the emergence of an active component in the ADP. The application of low-voltage-activated (LVA) calcium channel blockers such as nickel or mibefradil suppressed the generation of the active ADP component and burst-spiking behavior. In agreement with the development of the ADP waveform and burst-spiking behavior, voltage-clamp experiments in dissociated pyramidal neurons showed an increase in the LVA calcium current in P16-19 vs P9-12. Finally, we found that a reduction of extracellular calcium levels decreases the percentage of burst-spiking cells due to a reduction in the active component of the ADP. We conclude that a major contribution of LVA calcium channels to ADP determines the bursting capability of CA1 pyramidal neurons during a transitional postnatal period in contrast to adulthood.

  6. Severe acute respiratory syndrome coronavirus E protein transports calcium ions and activates the NLRP3 inflammasome.

    PubMed

    Nieto-Torres, Jose L; Verdiá-Báguena, Carmina; Jimenez-Guardeño, Jose M; Regla-Nava, Jose A; Castaño-Rodriguez, Carlos; Fernandez-Delgado, Raul; Torres, Jaume; Aguilella, Vicente M; Enjuanes, Luis

    2015-11-01

    Severe acute respiratory syndrome coronavirus (SARS-CoV) envelope (E) protein is a viroporin involved in virulence. E protein ion channel (IC) activity is specifically correlated with enhanced pulmonary damage, edema accumulation and death. IL-1β driven proinflammation is associated with those pathological signatures, however its link to IC activity remains unknown. In this report, we demonstrate that SARS-CoV E protein forms protein-lipid channels in ERGIC/Golgi membranes that are permeable to calcium ions, a highly relevant feature never reported before. Calcium ions together with pH modulated E protein pore charge and selectivity. Interestingly, E protein IC activity boosted the activation of the NLRP3 inflammasome, leading to IL-1β overproduction. Calcium transport through the E protein IC was the main trigger of this process. These findings strikingly link SARS-CoV E protein IC induced ionic disturbances at the cell level to immunopathological consequences and disease worsening in the infected organism.

  7. Endoplasmic reticulum potassium-hydrogen exchanger and small conductance calcium-activated potassium channel activities are essential for ER calcium uptake in neurons and cardiomyocytes.

    PubMed

    Kuum, Malle; Veksler, Vladimir; Liiv, Joanna; Ventura-Clapier, Renee; Kaasik, Allen

    2012-02-01

    Calcium pumping into the endoplasmic reticulum (ER) lumen is thought to be coupled to a countertransport of protons through sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) and the members of the ClC family of chloride channels. However, pH in the ER lumen remains neutral, which suggests a mechanism responsible for proton re-entry. We studied whether cation-proton exchangers could act as routes for such a re-entry. ER Ca(2+) uptake was measured in permeabilized immortalized hypothalamic neurons, primary rat cortical neurons and mouse cardiac fibers. Replacement of K(+) in the uptake solution with Na(+) or tetraethylammonium led to a strong inhibition of Ca(2+) uptake in neurons and cardiomyocytes. Furthermore, inhibitors of the potassium-proton exchanger (quinine or propranolol) but not of the sodium-proton exchanger reduced ER Ca(2+) uptake by 56-82%. Externally added nigericin, a potassium-proton exchanger, attenuated the inhibitory effect of propranolol. Inhibitors of small conductance calcium-sensitive K(+) (SK(Ca)) channels (UCL 1684, dequalinium) blocked the uptake of Ca(2+) by the ER in all preparations by 48-94%, whereas inhibitors of other K(+) channels (IK(Ca), BK(Ca) and K(ATP)) had no effect. Fluorescence microscopy and western blot analysis revealed the presence of both SK(Ca) channels and the potassium-proton exchanger leucine zipper-EF-hand-containing transmembrane protein 1 (LETM1) in ER in situ and in the purified ER fraction. The data obtained demonstrate that SK(Ca) channels and LETM1 reside in the ER membrane and that their activity is essential for ER Ca(2+) uptake.

  8. Comparative study of calcium alginate, activated carbon, and their composite beads on methylene blue adsorption.

    PubMed

    Hassan, A F; Abdel-Mohsen, A M; Fouda, Moustafa M G

    2014-02-15

    Three adsorbents, calcium alginate beads (AB), sodium hydroxide activated carbon based coconut shells (C), and calcium alginate/activated carbon composite beads (ACB) were prepared. Their textural properties were characterized by N2-adsorption at -196°C and scanning electron microscopy. The porosity, surface area and total pore volume of C>ACB>AB, but AB adsorbent was more acidic function groups more than the other adsorbents. Adsorption experiments were conducted to examine the effects of adsorbent dosage, pH, time, temperature and initial concentration of methylene blue. Methylene blue adsorption on C, AB and ACB was observed at pH>6 to avoid the competition of H(+). The amount of dye adsorbed increases as the adsorbent dosage increase. Adsorption of dye follows pseudo-second order mechanism. Thermodynamic studies show spontaneous and endothermic nature of the overall adsorption process.

  9. Active and passive calcium transport systems in plant cells. Progress report, May 1986--January 1991

    SciTech Connect

    Sze, H.

    1991-12-31

    The ability to change cytoplasmic Ca{sup 2+} levels ([Ca{sup 2+}]) by cells has made this cation a key regulator of many biological processes. Cytoplasmic [Ca{sup 2+}] is determined by the coordination of passive Ca{sup 2+} fluxes which increase cytosolic [Ca{sup 2+}] and active Ca{sup 2+} transport systems that lower cytosolic [Ca{sup 2+}]. The mechanisms by which plant cells achieve this is poorly understood. We have initially used isolated vesicles from the plasma membrane or organellar membranes to study Ca{sup 2+} transport systems in oat roots (a monocot) and carrot suspension cells (a dicot). The objectives of the proposal were to identify and characterize active (energy-dependent) and passive calcium transport systems that work together to regulate calcium levels in the cytoplasm of plant cells.

  10. Molecular heterogeneity of large-conductance calcium-activated potassium channels in canine intracardiac ganglia.

    PubMed

    Selga, Elisabet; Pérez-Serra, Alexandra; Moreno-Asso, Alba; Anderson, Seth; Thomas, Kristen; Desai, Mayurika; Brugada, Ramon; Pérez, Guillermo J; Scornik, Fabiana S

    2013-01-01

    Large conductance calcium-activated potassium (BK) channels are widely expressed in the nervous system. We have recently shown that principal neurons from canine intracardiac ganglia (ICG) express a paxilline- and TEA-sensitive BK current, which increases neuronal excitability. In the present work, we further explore the molecular constituents of the BK current in canine ICG. We found that the β1 and β4 regulatory subunits are expressed in ICG. Single channel voltage-dependence at different calcium concentrations suggested that association of the BKα with a particular β subunit was not enough to explain the channel activity in this tissue. Indeed, we detected the presence of several splice variants of the BKα subunit. In conclusion, BK channels in canine ICG may result from the arrangement of different BKα splice variants, plus accessory β subunits. The particular combinations expressed in canine IC neurons likely rule the excitatory role of BK current in this tissue.

  11. Calcium-Activated Potassium Channels at Nodes of Ranvier Secure Axonal Spike Propagation.

    PubMed

    Gründemann, Jan; Clark, Beverley A

    2015-09-22

    Functional connectivity between brain regions relies on long-range signaling by myelinated axons. This is secured by saltatory action potential propagation that depends fundamentally on sodium channel availability at nodes of Ranvier. Although various potassium channel types have been anatomically localized to myelinated axons in the brain, direct evidence for their functional recruitment in maintaining node excitability is scarce. Cerebellar Purkinje cells provide continuous input to their targets in the cerebellar nuclei, reliably transmitting axonal spikes over a wide range of rates, requiring a constantly available pool of nodal sodium channels. We show that the recruitment of calcium-activated potassium channels (IK, K(Ca)3.1) by local, activity-dependent calcium (Ca(2+)) influx at nodes of Ranvier via a T-type voltage-gated Ca(2+) current provides a powerful mechanism that likely opposes depolarizing block at the nodes and is thus pivotal to securing continuous axonal spike propagation in spontaneously firing Purkinje cells.

  12. Calcium-Activated Potassium Channels at Nodes of Ranvier Secure Axonal Spike Propagation

    PubMed Central

    Gründemann, Jan; Clark, Beverley A.

    2015-01-01

    Summary Functional connectivity between brain regions relies on long-range signaling by myelinated axons. This is secured by saltatory action potential propagation that depends fundamentally on sodium channel availability at nodes of Ranvier. Although various potassium channel types have been anatomically localized to myelinated axons in the brain, direct evidence for their functional recruitment in maintaining node excitability is scarce. Cerebellar Purkinje cells provide continuous input to their targets in the cerebellar nuclei, reliably transmitting axonal spikes over a wide range of rates, requiring a constantly available pool of nodal sodium channels. We show that the recruitment of calcium-activated potassium channels (IK, KCa3.1) by local, activity-dependent calcium (Ca2+) influx at nodes of Ranvier via a T-type voltage-gated Ca2+ current provides a powerful mechanism that likely opposes depolarizing block at the nodes and is thus pivotal to securing continuous axonal spike propagation in spontaneously firing Purkinje cells. PMID:26344775

  13. Calcium mobilization is both required and sufficient for initiating chromatin decondensation during activation of peripheral T-cells

    PubMed Central

    Lee, Megan D.; Bingham, Kellie N.; Mitchell, Taylor Y.; Meredith, Jenna L.; Rawlings, Jason S.

    2014-01-01

    Antigen engagement of the T-cell receptor (TCR) induces a rapid and dramatic decondensation of chromatin that is necessary for T-cell activation. This decondensation makes T-cells competent to respond to Interleukin-2 providing a mechanism to ensure clonotypic proliferation during an immune response. Using murine T-cells, we investigated the mechanism by which TCR signaling can initiate chromatin decondensation, focusing on the role of calcium mobilization. During T-cell activation, calcium is first released from intracellular stores, followed by influx of extracellular calcium via store operated calcium entry. We show that mobilization of intracellular calcium is required for TCR-induced chromatin decondensation. However, the decondensation is not dependent on the activity of the downstream transcription factor NFAT. Furthermore, we show that the influx of extracellular calcium is dispensable for initiating chromatin decondensation. Finally, we show that mobilization of calcium from intracellular stores is sufficient to induce decondensation, independent of TCR engagement. Collectively, our data suggest that chromatin decondensation in peripheral T-cells is controlled by modulating intracellular calcium levels. PMID:25453467

  14. Calcium transients in asymmetrically activated skeletal muscle fibers.

    PubMed Central

    Trube, G; Lopez, J R; Taylor, S R

    1981-01-01

    Skeletal muscle fibers of the frog Rana temporaria were held just taut and stimulated transversely by unidirectional electrical fields. We observed the reversible effects of stimulus duration (0.1-100 ms) and strength on action potentials, intracellular Ca2+ transients (monitored by aequorin), and contractile force during fixed-end contractions. Long duration stimuli (e.g., 10 ms) induced a maintained depolarization on the cathodal side of a cell and a maintained hyperpolarization on its anodal side. The hyperpolarization of the side facing the anode prevented the action potential from reaching mechanical threshold during strong stimuli. Variation of the duration or strength of a stimulus changed the luminescent response from a fiber injected with aequorin. Thus, the intracellular Ca2+ released during excitation-contraction coupling could be changed by the stimulus parameters. Prolongation of a stimulus at field strengths above 1.1 x rheobase decreased the amplitude of aequorin signals and the force of contractions. The decreases in aequorin and force signals from a given fiber paralleled one another and depended on the stimulus strength, but not on the stimulus polarity. These changes were completely reversible for stimulus strengths up to at least 4.2 x rheobase. The graded decreases in membrane depolarization, aequorin signals, and contractile force were correlated with the previously described folding of myofibrils in fibers allowed to shorten in response to the application of a long duration stimulus. The changes in aequorin signals and force suggest an absence of myofilament activation by Ca2+ in the section of the fiber closest to the anode. The results imply that injected aequorin distributes circumferentially in frog muscle with a coefficient of at least 10(-7) cm2/s, which is not remarkably different from the previously measured coefficient of 5 x 10(-8) cm2/s for its diffusion lengthwise. PMID:6976801

  15. Thermal Decomposition of Calcium Perchlorate/Iron-Mineral Mixtures: Implications of the Evolved Oxygen from the Rocknest Eolian Deposit in Gale Crater, Mars

    NASA Technical Reports Server (NTRS)

    Bruck, A. M.; Sutter, B.; Ming, D. W.; Mahaffy, P.

    2014-01-01

    A major oxygen release between 300 and 500 C was detected by the Mars Curiosity Rover Sample Analysis at Mars (SAM) instrument at the Rocknest eolian deposit. Thermal decomposition of perchlorate (ClO4-) salts in the Rocknest samples are a possible explanation for this evolved oxygen release. Releative to Na-, K-, Mg-, and Fe-perchlorate, the thermal decomposition of Ca-perchlorate in laboratory experiments released O2 in the temperature range (400-500degC) closest to the O2 release temperatures observed for the Rocknest material. Furthermore, calcium perchlorate could have been the source of Cl in the chlorinated-hydrocarbons species that were detected by SAM. Different components in the Martian soil could affect the decomposition temperature of calcium per-chlorate or another oxychlorine species. This interaction of the two components in the soil could result in O2 release temperatures consistent with those detected by SAM in the Rocknest materials. The decomposition temperatures of various alkali metal perchlorates are known to decrease in the presence of a catalyst. The objective of this work is to investigate catalytic interactions on calcium perchlorate from various iron-bearing minerals known to be present in the Rocknest material

  16. Characterization of Beta-leptinotarsin-h and the Effects of Calcium Flux Antagonists on its Activity

    DTIC Science & Technology

    2007-11-02

    observation eliminated non-selective cation channels such as nicotinic, glutamatergic, purinergic P2X, and serotoni- nergic 5 - HT3 ligand-operated Ca 2C...Characterization of b-leptinotarsin-h and the effects of calcium flux antagonists on its activity Richard D. Croslanda,*, Richard W. Fitchb,1, Harry...presynaptic nerve terminals. We tested antagonists of Ca2C flux for their effects on b-leptinotarsin-h-stimulated Ca2C uptake in rat brain synaptosomes

  17. Control of Spontaneous Firing Patterns by the Selective Coupling of Calcium Currents to Calcium Activated Potassium Currents in Striatal Cholinergic Interneurons

    PubMed Central

    Goldberg, Joshua A.; Wilson, Charles J.

    2005-01-01

    The spontaneous firing patterns of striatal cholinergic interneurons are sculpted by potassium currents that give rise to prominent afterhyperpolarizations (AHPs): BK currents contribute to action potential (AP) repolarization; SK currents generate an apamin-sensitive medium AHP (mAHP) following each AP; and bursts of APs generate long-lasting slow AHPs (sAHPs) due to apamin-insensitive currents. As all these currents are calcium-dependent, we conducted voltage- and current-clamp whole-cell recordings while pharmacologically manipulating calcium channels of the plasma membrane and intracellular stores to determine what sources of calcium activate the currents underlying AP repolarization and the AHPs. The Cav2.2 (N type) blocker, omega-conotoxin GVIA (1 μM), was the only blocker that significantly reduced the mAHP, and it induced a transition to rhythmic bursting in a third of the cells tested. Cav1 (L type) blockers (10 μM dihydropyridines) were the only ones that significantly reduced the sAHP. When applied to cells induced to burst with apamin, dihydropyridines reduced the sAHPs and abolished bursting. Depletion of intracellular stores with 10 mM caffeine also significantly reduced the sAHP current, and reversibly regularized firing. Application of 1 μM omega-conotoxin MVIIC (a Cav2.1/2.2 blocker) broadened APs, but had a negligible effect on APs in cells whose BK channels were already blocked by submillimolar tetraethylammonium chloride, indicating that Cav2.1 (Q type) channels provide the calcium to activate BK channels that repolarize the AP. Thus, calcium currents are selectively coupled to the calcium-dependent potassium currents underlying the AHPs thereby creating mechanisms for control of these neurons’ spontaneous firing patterns. PMID:16267230

  18. Optical imaging of neuronal activity in tissue labeled by retrograde transport of Calcium Green Dextran.

    PubMed

    McPherson, D R; McClellan, A D; O'Donovan, M J

    1997-05-01

    In many neurophysiological studies it is desirable to simultaneously record the activity of a large number of neurons. This is particularly true in the study of vertebrate motor systems that generate rhythmic behaviors, such as the pattern generator for locomotion in vertebrate spinal cord. Optical imaging of neurons labeled with appropriate fluorescent dyes, in which fluorescence is activity-dependent, provides a means to record the activity of many neurons at the same time, while also providing fine spatial resolution of the position and morphology of active neurons. Voltage-sensitive dyes have been explored for this purpose and have the advantage of rapid response to transmembrane voltage changes. However, voltage-sensitive dyes bleach readily, which results in phototoxic damage and limits the time that labeled neurons can be imaged. In addition, the signal-to-noise ratio is typically small, so that averaging of responses is usually required. As an alternative to voltage-sensitive dyes, calcium-sensitive dyes can exhibit large changes in fluorescence. Most neurons contain voltage-sensitive Ca2+ channels, and numerous reports indicate that neuronal activity is accompanied by increased intracellular Ca2+ concentration. In this protocol we describe a method to use retrograde transport of the dextran conjugate of a calcium-sensitive dye (Calcium Green Dextran) to label selectively populations of brain and spinal interneurons in a primitive vertebrate (lamprey), for subsequent video-rate imaging of changes in intracellular fluorescence during neuronal activity. Although described with specific reference to lampreys, the technique has also been applied to embryonic chick spinal cord and larval zebrafish preparations and should be easily adaptable to other systems. The most significant novel feature of the protocol is the use of retrograde axonal transport to selectively fill neurons that have known axonal trajectories. Using lampreys, we have obtained activity

  19. Macrophage-mediated osteogenesis activation in co-culture with osteoblast on calcium silicate cement.

    PubMed

    Tu, Ming-Gene; Chen, Yi-Wen; Shie, Ming-You

    2015-12-01

    The use of calcium silicate (CS) cement holds great promise for bone substitute biomaterials. However, the effects of CS on osteoblast and macrophage cells are not fully understood. This study examines cell proliferation and differentiation of mono- or co-cultured MC3T3-E1 and Raw 264.7 cells on CS cement. Very few studies to date have looked at the effects of osteoblast and macrophages on biomaterial-regulated osteogenesis. In this study the proliferation and differentiation of MC3T3-E1, Raw 264.7 and co-cultured MC3T3-E1/Raw 264.7 on CS cements have been analyzed using a PrestoBlue kit and ELISA. In addition, the effect of macrophages on CS-coordinated osteogenesis of MC3T3-E1 has been investigated. Results show that MC3T3-E1, Raw 264.7 and co-cultured MC3T3-E1/Raw 264.7 adhere to and proliferate well on the CS cement. In a co-culture, the CS cements inhibit receptor activator of nuclear factor kappa B ligand expression of both genes and proteins in Raw 264.7 cells when compared to those grown in mono-cultured system. Ca deposition of MC3T3-E1 in the co-culture is higher than that of cells in a mono-culture. Bone morphogenetic protein 2 (BMP2) is also significantly up-regulated by the CS cement stimulation, indicating that macrophages may participate in the CS stimulated osteogenesis. Interestingly, when macrophage are cultured with BMP2 receptor-blocking MC3T3-E1 on the CS cements, the osteogenesis differentiation of the cells is significantly inhibited, indicating the important role of macrophages in biomaterial-induced osteogenesis via BMP2 receptors. It is assumed that it is an increase in the secretion of the BMP2 from the Raw 264.7 cell that is primarily involved in the promotion of the osteogenesis of the MC3T3-E1. These results provide valuable insights into both the mechanism of CS-stimulated osteogenesis, and strategies to optimize the evaluation system for the in vitro osteogenesis capacity of bone substitute biomaterials.

  20. Modulation of intracellular calcium concentrations and T cell activation by prickly pear polyphenols.

    PubMed

    Aires, Virginie; Adote, Sylvie; Hichami, Aziz; Moutairou, Kabirou; Boustani, Es-Saddik E; Khan, Naim A

    2004-05-01

    Opuntia ficus indica (prickly pear) polyphenolic compounds (OFPC) triggered an increase in [Ca2+]i in human Jurkat T-cell lines. Furthermore, OFPC-induced rise in [Ca2+]i was significantly curtailed in calcium-free buffer (0% Ca2+) as compared to that in 100% Ca2+ medium. Preincubation of cells with tyrphostin A9, an inhibitor of Ca2+ release-activated Ca2+ (CRAC) channels, significantly diminished the OFPC-induced sustained response on the increases in [Ca2+]i. Lanthanum and nifedipine, the respective inhibitors of voltage-dependent and L-type calcium channels, failed to curtail significantly the OFPC-induced calcium response. As OFPC still stimulated increases in [Ca2+]i in 0% Ca2+ medium, the role of intracellular calcium was investigated. Hence, addition of thapsigargin (TG), an inhibitor of Ca2+-ATPase of the endoplasmic reticulum (ER), during the OFPC-induced peak response exerted an additive effect, indicating that the mechanism of action of these two agents are different. Furthermore, U73122, an inhibitor of IP3 production, completely abolished increases in [Ca2+]i, induced by OFPC, suggesting that these polyphenols induce the production of IP3 that recruits calcium from ER pool. Polyphenolic compounds do act extracellularly as addition of fatty acid-free bovine serum albumin (BSA) significantly diminished the rise in [Ca2+]i evoked by the formers. OFPC also induced plasma membrane hyperpolarisation which was reversed by addition of BSA. OFPC were found to curtail the expression of IL-2 mRNA and T-cell blastogenesis. Together these results suggest that OFPC induce increases in [Ca2+]i via ER pool and opening of CRAC channels, and exert immunosuppressive effects in Jurkat T-cells.

  1. Sucrose increases calcium-dependent protein kinase and phosphatase activities in potato plants.

    PubMed

    Raíces, M; MacIntosh, G C; Ulloa, R M; Gargantini, P R; Vozza, N F; Téllez-Inón, M T

    2003-09-01

    The effect of sucrose on tuber formation, calcium-dependent protein kinase (CDPK) and phosphatase activities was analysed using in vitro cultured potato plants. In short treatments, sucrose induced CDPK and phosphatase activities. In long treatments, sucrose induced tuber formation in the absence of other tuber inducing stimuli. Sorbitol caused a minor increase in CDPK activity and affected plant morphology but did not induce tuber development. The addition of the protein kinase inhibitor Staurosporine precluded sucrose-induced tuberization. Altogether, our results suggest that phosphorylation/dephosphorylation events are involved in sucrose-induced tuber development.

  2. Calcium-dependent depletion zones in the cortical microtubule array coincide with sites of, but do not regulate, wall ingrowth papillae deposition in epidermal transfer cells

    PubMed Central

    Zhang, Hui-ming; Talbot, Mark J.; McCurdy, David W.; Patrick, John W.; Offler, Christina E.

    2015-01-01

    Trans-differentiation to a transfer-cell morphology is characterized by the localized deposition of wall ingrowth papillae that protrude into the cytosol. Whether the cortical microtubule array directs wall ingrowth papillae formation was investigated using a Vicia faba cotyledon culture system in which their adaxial epidermal cells were spontaneously induced to trans-differentiate to transfer cells. During deposition of wall ingrowth papillae, the aligned cortical microtubule arrays in precursor epidermal cells were reorganized into a randomized array characterized by circular depletion zones. Concurrence of the temporal appearance, spatial pattern, and size of depletion zones and wall ingrowth papillae was consistent with each papilla occupying a depletion zone. Surprisingly, microtubules appeared not to regulate construction of wall ingrowth papillae, as neither depolymerization nor stabilization of cortical microtubules changed their deposition pattern or morphology. Moreover, the size and spatial pattern of depletion zones was unaltered when the formation of wall ingrowth papillae was blocked by inhibiting cellulose biosynthesis. In contrast, the depletion zones were absent when the cytosolic calcium plumes, responsible for directing wall ingrowth papillae formation, were blocked or dissipated. Thus, we conclude that the depletion zones within the cortical microtubule array result from localized depolymerization of microtubules initiated by elevated cytosolic Ca2+ levels at loci where wall ingrowth papillae are deposited. The physiological significance of the depletion zones as a mechanism to accommodate the construction of wall ingrowth papillae without compromising maintenance of the plasma membrane–microtubule inter-relationship is discussed. PMID:26136268

  3. Antiresorption implant coatings based on calcium alendronate and octacalcium phosphate deposited by matrix assisted pulsed laser evaporation.

    PubMed

    Boanini, Elisa; Torricelli, Paola; Forte, Lucia; Pagani, Stefania; Mihailescu, Natalia; Ristoscu, Carmen; Mihailescu, Ion N; Bigi, Adriana

    2015-12-01

    The integration of an implant material with bone tissue depends on the chemistry and physics of the implant surface. In this study we applied matrix assisted pulsed laser evaporation (MAPLE) in order to synthesize calcium alendronate monohydrate (a bisphosphonate obtained by calcium sequestration from octacalcium phosphate by alendronate) and calcium alendronate monohydrate/octacalcium phosphate composite thin films on titanium substrates. Octacalcium phosphate coatings were prepared as reference material. The powders, which were synthesized in aqueous medium, were suspended in deionised water, frozen at liquid nitrogen temperature and used as targets for MAPLE experiments. The transfer was conducted with a KrF* excimer laser source (λ = 248 nm, τFWHM ≤ 25 ns) in mild conditions of temperature and pressure. XRD, FTIR and SEM analyses confirmed that the coatings contain the same crystalline phases as the as-prepared powder samples. Osteoblast derived from stem cells and osteoclast derived from monocytes of osteoporotic subjects were co-cultured on the coatings up to 14 days. Osteoclast displayed significantly reduced proliferation and differentiation in the presence of calcium alendronate monohydrate, pointing to a clear role of the coatings containing this bisphosphonate on inhibiting excessive bone resorption. At variance, osteoblast production of alkaline phosphatase and type I pro-collagen were promoted by the presence of bisphosphonate, which also decreased the production of interleukin 6. The positive influence towards osteoblast differentiation was even more enhanced in the composite coatings, thanks to the presence of octacalcium phosphate.

  4. The longitudinal effects of physical activity and dietary calcium on bone mass accrual across stages of pubertal development.

    PubMed

    Lappe, Joan M; Watson, Patrice; Gilsanz, Vicente; Hangartner, Thomas; Kalkwarf, Heidi J; Oberfield, Sharon; Shepherd, John; Winer, Karen K; Zemel, Babette

    2015-01-01

    Childhood and adolescence are critical periods of bone mineral content (BMC) accrual that may have long-term consequences for osteoporosis in adulthood. Adequate dietary calcium intake and weight-bearing physical activity are important for maximizing BMC accrual. However, the relative effects of physical activity and dietary calcium on BMC accrual throughout the continuum of pubertal development in childhood remains unclear. The purpose of this study was to determine the effects of self-reported dietary calcium intake and weight-bearing physical activity on bone mass accrual across the five stages of pubertal development in a large, diverse cohort of US children and adolescents. The Bone Mineral Density in Childhood study was a mixed longitudinal study with 7393 observations on 1743 subjects. Annually, we measured BMC by dual-energy X-ray absorptiometry (DXA), physical activity and calcium intake by questionnaire, and pubertal development (Tanner stage) by examination for up to 7 years. Mixed-effects regression models were used to assess physical activity and calcium intake effects on BMC accrual at each Tanner stage. We found that self-reported weight-bearing physical activity contributed to significantly greater BMC accrual in both sexes and racial subgroups (black and nonblack). In nonblack males, the magnitude of the activity effect on total body BMC accrual varied among Tanner stages after adjustment for calcium intake; the greatest difference between high- and low-activity boys was in Tanner stage 3. Calcium intake had a significant effect on bone accrual only in nonblack girls. This effect was not significantly different among Tanner stages. Our findings do not support differential effects of physical activity or calcium intake on bone mass accrual according to maturational stage. The study demonstrated significant longitudinal effects of weight-bearing physical activity on bone mass accrual through all stages of pubertal development.

  5. Diffusion and extrusion shape standing calcium gradients during ongoing parallel fiber activity in dendrites of Purkinje neurons.

    PubMed

    Schmidt, Hartmut; Arendt, Oliver; Eilers, Jens

    2012-09-01

    Synaptically induced calcium transients in dendrites of Purkinje neurons (PNs) play a key role in the induction of plasticity in the cerebellar cortex (Ito, Physiol Rev 81:1143-1195, 2001). Long-term depression at parallel fiber-PN synapses can be induced by stimulation paradigms that are associated with long-lasting (>1 min) calcium signals. These signals remain strictly localized (Eilers et al., Learn Mem 3:159-168, 1997), an observation that was rather unexpected, given the high concentration of the mobile endogenous calcium-binding proteins parvalbumin and calbindin in PNs (Fierro and Llano, J Physiol (Lond) 496:617-625, 1996; Kosaka et al., Exp Brain Res 93:483-491, 1993). By combining two-photon calcium imaging experiments in acute slices with numerical computer simulations, we found that significant calcium diffusion out of active branches indeed takes places. It is outweighed, however, by rapid and powerful calcium extrusion along the dendritic shaft. The close interplay of diffusion and extrusion defines the spread of calcium between active and inactive dendritic branches, forming a steep gradient in calcium with drop ranges of ~13 μm (interquartile range, 10-18 μm).

  6. Melanocortin 4 receptor activation inhibits presynaptic N-type calcium channels in amygdaloid complex neurons.

    PubMed

    Agosti, Francina; López Soto, Eduardo J; Cabral, Agustina; Castrogiovanni, Daniel; Schioth, Helgi B; Perelló, Mario; Raingo, Jesica

    2014-09-01

    The melanocortin 4 receptor (MC4R) is a G protein-coupled receptor involved in food intake and energy expenditure regulation. MC4R activation modifies neuronal activity but the molecular mechanisms by which this regulation occurs remain unclear. Here, we tested the hypothesis that MC4R activation regulates the activity of voltage-gated calcium channels and, as a consequence, synaptic activity. We also tested whether the proposed effect occurs in the amygdala, a brain area known to mediate the anorexigenic actions of MC4R signaling. Using the patch-clamp technique, we found that the activation of MC4R with its agonist melanotan II specifically inhibited 34.5 ± 1.5% of N-type calcium currents in transiently transfected HEK293 cells. This inhibition was concentration-dependent, voltage-independent and occluded by the Gαs pathway inhibitor cholera toxin. Moreover, we found that melanotan II specifically inhibited 25.9 ± 2.0% of native N-type calcium currents and 55.4 ± 14.4% of evoked inhibitory postsynaptic currents in mouse cultured amygdala neurons. In vivo, we found that the MC4R agonist RO27-3225 increased the marker of cellular activity c-Fos in several components of the amygdala, whereas the N-type channel blocker ω conotoxin GVIA increased c-Fos expression exclusively in the central subdivision of the amygdala. Thus, MC4R specifically inhibited the presynaptic N-type channel subtype, and this inhibition may be important for the effects of melanocortin in the central subdivision of the amygdala.

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

    SciTech Connect

    Feng, Shan-Li; Sun, Ming-Rui; Li, Ting-Ting; Yin, Xin; Xu, Chang-Qing; Sun, Yi-Hua

    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

  8. Plasma membrane calcium flux, protein kinase C activation and smooth muscle contraction.

    PubMed

    Forder, J; Scriabine, A; Rasmussen, H

    1985-11-01

    Isolated perfused rabbit ear arteries contract when treated with 12-O-tetradecanoylphorbol-13-acetate (TPA), an activator of the calcium-activated, phospholipid-dependent protein kinase or C-kinase. Under conditions where the calcium concentration in the perfusate is 1.5 mM and the potassium concentration is 4.8 mM, there is a latent period of 70 +/- 19 min (mean +/- S.E.M., n = 10) between TPA addition and the onset of the contractile response. Once initiated, the contractile response is progressive and sustained. When perfusion conditions are altered in such a way as to modify calcium flux across the plasma membrane (i.e., raising the extracellular calcium concentration to 2.5 mM Ca++, raising the extracellular potassium concentration to 10 mM, and/or preincubating the tissues in media containing 100 nM Bay K 8644, a potent calcium channel agonist), the latency period between TPA addition and initiation of the contractile response is significantly reduced (2.5 mM Ca++, 37 +/- 7 min; 10 mM K+ and 2.5 mM Ca++, 11 +/- 3 min; 100 nM Bay K 8644 and 1.5 mM Ca++, 20 +/- 7 min; 100 nM Bay K 8644 and 2.5 mM Ca2+, 8.5 +/- 1.7 min; 10 mM K+ and 100 nM Bay K 8644, 11 +/- 5 min). Likewise, the combination of 2.5 mM calcium, 100 nM Bay K 8644, and 3.3 microM ouabain results in a contractile response 4.5 +/- 2.0 min after TPA addition (means +/- S.E.M., n = 4). Control tissues (absence of TPA addition) run simultaneously show no contractile responses to the various Ca++ flux regulators even after 90 min of incubation.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Synthesis of a new series of 4-aryl-1,4-dihydropyridines with calcium channel blocking and vasodilatory activity.

    PubMed

    Jain, P; Narang, G; Jindal, D P; Bansal, R; Calle, C; Carron, R; Pemberton, K; Harvey, A L

    2006-05-01

    Synthesis of a new series of 4-aryl-1,4-dihydropyridines possessing potential calcium channel blocking activity along with good vasodilatory profile is reported. The compounds were synthesized using modified Hantzsch condensation of various aldehydes with methyl 3-aminocrotonate in the presence of a catalytic amount of trifluoroacetic acid and subsequent alkylation with various hydrochlorides of dialkylaminoalkyl chlorides. In vitro calcium channel blocking activity has been evaluated in cultures of neonatal rat cortical neurons by measuring the inhibitory response at L-type calcium channels activated by veratridine. Many compounds exhibited moderate to significant calcium channel blockade around 1 microM. The vasodilatory activity was assessed on isolated rat thoracic aortic rings precontracted by phenylephrine/KCl (30 mM). Most of the compounds produced a concentration-dependent inhibition of the contractile response.

  10. Evaluation of the calcium-antagonist, antidiarrhoeic and central nervous system activities of Baccharis serraefolia.

    PubMed

    Tortoriello, J; Aguilar-Santamaría, L

    1996-09-01

    Baccharis serraefolia is a widely used plant to treat diarrhoea in Mexican traditional medicine. Although the methanolic extract of this plant has shown an important dose-dependent spasmolytic activity, its underlying mechanism has not been studied. In the present work, the methanolic extract of B. serraefolia significantly delayed the onset of tonic seizures induced by strychnine and pentylenetetrazol; besides, it diminished the death rate and number of animals that exhibited convulsions. It produced potentiation of the hypnotic effect of pentobarbital. Oral administration produced an inhibition of gastrointestinal transit in mice as effective as that produced by loperamide. As to the effect on smooth muscles, the active extract produced an inhibition of contraction induced electrically, which could not be reversed by naloxone. The calcium concentration-contraction curve showed a rightward displacement when the extract was added to isolated guinea pig ileum depolarized with high K+ and cumulative concentrations of Ca2+. The results suggest that the methanolic extract does not interact with classical opiate receptors and its effects, at least that produced on smooth muscle, may be due to a probable interference with calcium influx and/or calcium release from an intra-cellular store.

  11. Calcium activates the light-dependent conductance in melanopsin-expressing photoreceptors of amphioxus.

    PubMed

    Peinado, Gabriel; Osorno, Tomás; Gomez, María del Pilar; Nasi, Enrico

    2015-06-23

    Melanopsin, the photopigment of the "circadian" receptors that regulate the biological clock and the pupillary reflex in mammals, is homologous to invertebrate rhodopsins. Evidence supporting the involvement of phosphoinositides in light-signaling has been garnered, but the downstream effectors that control the light-dependent conductance remain unknown. Microvillar photoreceptors of the primitive chordate amphioxus also express melanopsin and transduce light via phospholipase-C, apparently not acting through diacylglycerol. We therefore examined the role of calcium in activating the photoconductance, using simultaneous, high time-resolution measurements of membrane current and Ca(2+) fluorescence. The light-induced calcium rise precedes the onset of the photocurrent, making it a candidate in the activation chain. Moreover, photolysis of caged Ca elicits an inward current of similar size, time course and pharmacology as the physiological photoresponse, but with a much shorter latency. Internally released calcium thus emerges as a key messenger to trigger the opening of light-dependent channels in melanopsin-expressing microvillar photoreceptors of early chordates.

  12. Development of a glial network in the olfactory nerve: role of calcium and neuronal activity.

    PubMed

    Koussa, Mounir A; Tolbert, Leslie P; Oland, Lynne A

    2010-11-01

    In adult olfactory nerves of mammals and moths, a network of glial cells ensheathes small bundles of olfactory receptor axons. In the developing antennal nerve (AN) of the moth Manduca sexta, the axons of olfactory receptor neurons (ORNs) migrate from the olfactory sensory epithelium toward the antennal lobe. Here we explore developmental interactions between ORN axons and AN glial cells. During early stages in AN glial-cell migration, glial cells are highly dye coupled, dividing glia are readily found in the nerve and AN glial cells label strongly for glutamine synthetase. By the end of this period, dye-coupling is rare, glial proliferation has ceased, glutamine synthetase labeling is absent, and glial processes have begun to extend to enwrap bundles of axons, a process that continues throughout the remainder of metamorphic development. Whole-cell and perforated-patch recordings in vivo from AN glia at different stages of network formation revealed two potassium currents and an R-like calcium current. Chronic in vivo exposure to the R-type channel blocker SNX-482 halted or greatly reduced AN glial migration. Chronically blocking spontaneous Na-dependent activity by injection of tetrodotoxin reduced the glial calcium current implicating an activity-dependent interaction between ORNs and glial cells in the development of glial calcium currents.

  13. Calcium activates the light-dependent conductance in melanopsin-expressing photoreceptors of amphioxus

    PubMed Central

    Peinado, Gabriel; Osorno, Tomás; Gomez, María del Pilar; Nasi, Enrico

    2015-01-01

    Melanopsin, the photopigment of the “circadian” receptors that regulate the biological clock and the pupillary reflex in mammals, is homologous to invertebrate rhodopsins. Evidence supporting the involvement of phosphoinositides in light-signaling has been garnered, but the downstream effectors that control the light-dependent conductance remain unknown. Microvillar photoreceptors of the primitive chordate amphioxus also express melanopsin and transduce light via phospholipase-C, apparently not acting through diacylglycerol. We therefore examined the role of calcium in activating the photoconductance, using simultaneous, high time-resolution measurements of membrane current and Ca2+ fluorescence. The light-induced calcium rise precedes the onset of the photocurrent, making it a candidate in the activation chain. Moreover, photolysis of caged Ca elicits an inward current of similar size, time course and pharmacology as the physiological photoresponse, but with a much shorter latency. Internally released calcium thus emerges as a key messenger to trigger the opening of light-dependent channels in melanopsin-expressing microvillar photoreceptors of early chordates. PMID:26056310

  14. Energetic basis of catalytic activity of layered nanophase calcium manganese oxides for water oxidation

    PubMed Central

    Birkner, Nancy; Nayeri, Sara; Pashaei, Babak; Najafpour, Mohammad Mahdi; Casey, William H.; Navrotsky, Alexandra

    2013-01-01

    Previous measurements show that calcium manganese oxide nanoparticles are better water oxidation catalysts than binary manganese oxides (Mn3O4, Mn2O3, and MnO2). The probable reasons for such enhancement involve a combination of factors: The calcium manganese oxide materials have a layered structure with considerable thermodynamic stability and a high surface area, their low surface energy suggests relatively loose binding of H2O on the internal and external surfaces, and they possess mixed-valent manganese with internal oxidation enthalpy independent of the Mn3+/Mn4+ ratio and much smaller in magnitude than the Mn2O3-MnO2 couple. These factors enhance catalytic ability by providing easy access for solutes and water to active sites and facile electron transfer between manganese in different oxidation states. PMID:23667149

  15. Calcium-sensing receptor activation in chronic kidney disease: effects beyond parathyroid hormone control.

    PubMed

    Massy, Ziad A; Hénaut, Lucie; Larsson, Tobias E; Vervloet, Marc G

    2014-11-01

    Secondary hyperparathyroidism (SHPT) is an important complication of advanced chronic kidney disease (CKD). Cinacalcet, an allosteric modulator of the calcium-sensing receptor (CaSR) expressed in parathyroid glands, is the only calcimimetic approved to treat SHPT in patients on dialysis. By enhancing CaSR sensitivity for plasma extracellular calcium (Ca(2+)0), cinacalcet reduces serum parathyroid hormone, Ca(2+)0, and serum inorganic phosphorous concentrations, allowing better control of SHPT and CKD-mineral and bone disorders. Of interest, the CaSR also is expressed in a variety of tissues where its activation regulates diverse cellular processes, including secretion, apoptosis, and proliferation. Thus, the existence of potential off-target effects of cinacalcet cannot be neglected. This review summarizes our current knowledge concerning the potential role(s) of the CaSR expressed in various tissues in CKD-related disorders, independently of parathyroid hormone control.

  16. T-type calcium channel blocker attenuates unilateral ureteral obstruction-induced renal interstitial fibrosis by activating the Nrf2 antioxidant pathway

    PubMed Central

    Chung, Sungjin; Kim, Soojeong; Kim, Minyoung; Koh, Eun Sil; Yoon, Hye Eun; Kim, Ho-Shik; Park, Cheol Whee; Chang, Yoon Sik; Shin, Seok Joon

    2016-01-01

    Besides its effect on high blood pressure, T-type calcium channel blocker is renoprotective in experimental models of renal fibrosis. However, the exact mechanism of T-type calcium channel blocker on tubulointerstitial fibrosis is unclear. We investigated whether the renoprotective effect of T-type calcium channel blocker is associated with modulation of the signaling of oxidative stress-induced renal fibrosis. Treatment with a non-hypotensive dose of efonidipine, a T-type calcium channel blocker, or nifedipine, an L-type channel blocker, was initiated one day before unilateral ureteral obstruction (UUO) in C57BL6/J mice, and was continued until 3 and 7 days after UUO. In the obstructed kidneys, treatment with efonidipine significantly attenuated interstitial fibrosis, collagen deposition and inflammation increased by UUO creation compared with treatment with nifedipine. Additionally, efonidipine significantly increased the expression of the antioxidant enzymes heme oxygenase-1, NAD(P)H: quinone oxidoreductase 1, catalase and superoxide dismutase 1. Increased apoptotic cell death and decreased B-cell lymphoma 2 expression were also significantly ameliorated by efonidipine. The expression of the histone acetyltransferase p300/CBP-associated factor, a regulator of inflammatory molecules, was significantly inhibited by efonidipine. These beneficial effects of efonipidine were attributed to the increased nuclear expression of nuclear factor-erythroid-2-related factor 2 (Nrf2) on UUO day 3 and the increased expressions of both total and nuclear Nrf2 with elevated Kelch-like ECH-associated protein 1 on UUO day 7. The data indicate that T-type calcium channel blocker exerts beneficial effects in renal interstitial fibrosis by activating Nrf2 and subsequent antioxidant enzymes. PMID:27904663

  17. The requirement of zinc and calcium ions for functional MMP activity in demineralized dentin matrices

    PubMed Central

    Tezvergil-Mutluay, Arzu; Agee, Kelli A.; Hoshika, Tomohiro; Carrilho, Marcela; Breschi, Lorenzo; Tjäderhane, Leo; Nishitani, Yoshihiro; Carvalho, Ricardo M.; Looney, Stephen; Tay, Franklin R.; Pashley, David H.

    2010-01-01

    The progressive degradation of resin-dentin bonds is due, in part, to the slow degradation of collagen fibrils in the hybrid layer by endogenous matrix metalloproteinases (MMPs) of the dentin matrix. In in vitro durability studies, the storage medium composition might be important because the optimum activity of MMPs requires both zinc and calcium. Objective This study evaluated the effect of different storage media on changes in matrix stiffness, loss of dry weight or solubilization of collagen from demineralized dentin beams incubated in vitro for up to 60 days. Methods Dentin beams (1×2×6mm) were completely demineralized in 10% phosphoric acid. After baseline measurements of dry mass and elastic modulus (E) (3-point bending, 15% strain) the beams were divided into 5 groups (n=11/group) and incubated at 37°C in either media containing both zinc and calcium designated as complete medium(CM), calcium-free medium, zinc-free medium, a doubled-zinc medium or water. Beams were retested at 3, 7, 14, 30, and 60 days of incubation. The incubation media was hydrolyzed with HCl for the quantitation of hydroxyproline (HOP) as an index of solubilization of collagen by MMPs. Data were analyzed using repeated measures of ANOVA. Results Both the storage medium and storage time showed significant effects on E, mass loss and HOP release (p<0.05). The incubation in CM resulted in relatively rapid and significant (p<0.05) decreases in stiffness, and increasing amounts of mass loss. The HOP content of the experimental media also increased with incubation time but was significantly lower (p<0.05) than in the control CM medium, the recommended storage medium. Conclusions The storage solutions used to age resin-dentin bonds should be buffered solutions that contain both calcium and zinc. The common use of water as an aging medium may underestimate the hydrolytic activity of endogenous dentin MMPs. PMID:20688380

  18. Preparation and fluorescence property of red-emitting Eu{sup 3+}-activated amorphous calcium silicate phosphor

    SciTech Connect

    Kojima, Yoshiyuki; Kamei, Shinnosuke; Nishimiya, Nobuyuki

    2010-02-15

    This paper describes the energy efficient synthesis of a red-emitting Eu{sup 3+}-activated amorphous calcium silicate phosphor produced by heating a Eu{sup 3+}-activated calcium silicate hydrate phosphor. Concentration quenching of the Eu{sup 3+}-activated calcium silicate hydrate phosphor was not observed and the emission intensity did not decrease up to a Eu/(Ca+Eu) atomic ratio of 0.46. Heating of the Eu{sup 3+}-activated calcium silicate hydrate (Eu/(Ca+Eu) atomic ratio = 0.32) phosphor produced an amorphous Eu{sup 3+}-activated calcium silicate phosphor, which had a maximum emission intensity at 870 {sup o}C and emitted in the red under near-ultraviolet irradiation (395 nm). The emission intensity of the Eu{sup 3+}-activated amorphous calcium silicate phosphor was about half that of a commercial BaMgAl{sub 10}O{sub 17}:Eu{sup 2+} phosphor, and shows great potential for application in white light-emitting diodes.

  19. Activation of the calcium sensing receptor with cinacalcet increases serum gastrin levels in healthy older subjects

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gastric acidity is postulated to enhance calcium absorption since calcium is better dissolved at low pH. Extracellular calcium stimulates gastrin and gastric acid secretion in humans. Ex vivo studies indicate that the calcium sensing receptor (CaR), which is expressed on the surface of human G cells...

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

    PubMed

    Zhang, Weiping; Schmelzeisen, Steffen; Parthier, Daniel; Frings, Stephan; Möhrlen, Frank

    2015-01-01

    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.

  1. The coexistence of geopolymeric gel and calcium silicate hydrate at the early stage of alkaline activation

    SciTech Connect

    Yip, C.K.; Lukey, G.C.; Deventer, J.S.J. van . E-mail: jannie@unimelb.edu.au

    2005-09-01

    Scanning electron microscopy was used to study the effects of the addition of ground granulated blast furnace slag (GGBFS) on the microstructure and mechanical properties of metakaolin (MK) based geopolymers. It was found that it is possible to have geopolymeric gel and calcium silicate hydrate (CSH) gel forming simultaneously within a single binder. The coexistence of these two phases is dependent on the alkalinity of the alkali activator and the MK / GGBFS mass ratio. It has been found that the formation of CSH gel together with the geopolymeric gel occurs only in a system at low alkalinity. In the presence of high concentrations of NaOH (> 7.5 M), the geopolymeric gel is the predominant phase formed with small calcium precipitates scattered within the binder. The coexistence of the two phases is not observed unless a substantial amount of a reactive calcium source is present initially. It is thought that voids and pores within the geopolymeric binder become filled with the CSH gel. This helps to bridge the gaps between the different hydrated phases and unreacted particles; thereby resulting in the observed increase in mechanical strength for these binders.

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

    PubMed Central

    Parthier, Daniel; Frings, Stephan; Möhrlen, Frank

    2015-01-01

    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. PMID:26558388

  3. Calcium uptake in rat liver mitochondria accompanied by activation of ATP-dependent potassium channel.

    PubMed

    Akopova, O V; Nosar, V I; Mankovskaya, I N; Sagach, V F

    2008-10-01

    The influence of potassium ions on calcium uptake in rat liver mitochondria is studied. It is shown that an increase in K+ and Ca2+ concentrations in the incubation medium leads to a decrease in calcium uptake in mitochondria together with a simultaneous increase in potassium uptake due to the potential-dependent transport of K+ in the mitochondrial matrix. Both effects are more pronounced in the presence of an ATP-dependent K+-channel (K+(ATP)-channel) opener, diazoxide (Dz). Activation of the K+(ATP)-channel by Dz alters the functional state of mitochondria and leads to an increase in the respiration rate in state 2 and a decrease in the oxygen uptake and the rate of ATP synthesis in state 3. The effect of Dz on oxygen consumption in state 3 is mimicked by valinomycin, but it is opposite to that of the classical protonophore uncoupler CCCP. It is concluded that the potential-dependent uptake of potassium is closely coupled to calcium transport and is an important parameter of energy coupling responsible for complex changes in oxygen consumption and Ca2+-transport properties of mitochondria.

  4. Internal calcium release and activation of sea urchin eggs by cGMP are independent of the phosphoinositide signaling pathway.

    PubMed Central

    Whalley, T; McDougall, A; Crossley, I; Swann, K; Whitaker, M

    1992-01-01

    We show that microinjecting cyclic GMP (cGMP) into unfertilized sea urchin eggs activates them by stimulating a rise in the intracellular free calcium ion concentration ([Ca2+]i). The increase in [Ca2+]i is similar in both magnitude and duration to the transient that activates the egg at fertilization. It is due to mobilization of calcium from intracellular stores but is not prevented by the inositol trisphosphate (InsP3) antagonist heparin. Furthermore, cGMP does not stimulate the eggs Na+/H+ antiport when the [Ca2+]i transient is blocked by the calcium chelator bis-(O-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA), suggesting that cGMP does not activate eggs by interacting with the their phosphoinositide signaling pathway. However, the [Ca2+]i increase and activation are prevented in eggs in which the InsP3-sensitive calcium stores have been emptied by the prior microinjection of the InsP3 analogue inositol 1,4,5-trisphosphorothioate. These data indicate that cGMP activates eggs by stimulating the release of calcium from an InsP3-sensitive calcium store via a novel, though unidentified, route independent of the InsP3 receptor. PMID:1320962

  5. Resveratrol suppresses calcium-mediated microglial activation and rescues hippocampal neurons of adult rats following acute bacterial meningitis.

    PubMed

    Sheu, Ji-Nan; Liao, Wen-Chieh; Wu, Un-In; Shyu, Ling-Yuh; Mai, Fu-Der; Chen, Li-You; Chen, Mei-Jung; Youn, Su-Chung; Chang, Hung-Ming

    2013-03-01

    Acute bacterial meningitis (ABM) is a serious disease with severe neurological sequelae. The intense calcium-mediated microglial activation and subsequently pro-inflammatory cytokine release plays an important role in eliciting ABM-related oxidative damage. Considering resveratrol possesses significant anti-inflammatory and anti-oxidative properties, the present study aims to determine whether resveratrol would exert beneficial effects on hippocampal neurons following ABM. ABM was induced by inoculating Klebsiella pneumoniae into adult rats intraventricularly. The time-of-flight secondary ion mass spectrometry (TOF-SIMS), Griffonia simplicifolia isolectin-B4 (GSA-IB4) and ionized calcium binding adaptor molecule 1 (Iba1) immunohistochemistry, enzyme-linked immunosorbent assay as well as malondialdehyde (MDA) measurement were used to examine the calcium expression, microglial activation, pro-inflammatory cytokine level, and extent of oxidative stress, respectively. In ABM rats, strong calcium signaling associated with enhanced microglial activation was observed in hippocampus. Increased microglial expression was coincided with intense production of pro-inflammatory cytokines and oxidative damage. However, in rats receiving resveratrol after ABM, the calcium intensity, microglial activation, pro-inflammatory cytokine and MDA levels were all significantly decreased. Quantitative data showed that much more hippocampal neurons were survived in resveratrol-treated rats following ABM. As resveratrol successfully rescues hippocampal neurons from ABM by suppressing the calcium-mediated microglial activation, therapeutic use of resveratrol may act as a promising strategy to counteract the ABM-induced neurological damage.

  6. Activity-dependent regulation of calcium and ribosomes in the chick cochlear nucleus.

    PubMed

    Call, C L; Hyson, R L

    2016-03-01

    Cochlea removal results in the death of 20-30% of neurons in the chick cochlear nucleus, nucleus magnocellularis (NM). Two potentially cytotoxic events, a dramatic rise in intracellular calcium concentration ([Ca(2+)]i) and a decline in the integrity of ribosomes are observed within 1h of deafferentation. Glutamatergic input from the auditory nerve has been shown to preserve NM neuron health by activating metabotropic glutamate receptors (mGluRs), maintaining both normal [Ca(2+)]i and ribosomal integrity. One interpretation of these results is that a common mGluR-activated signaling cascade is required for the maintenance of both [Ca(2+)]i and ribosomal integrity. This could happen if both responses are influenced directly by a common messenger, or if the loss of mGluR activation causes changes in one component that secondarily causes changes in the other. The present studies tested this common-mediator hypothesis in slice preparations by examining activity-dependent regulation of [Ca(2+)]i and ribosomes in the same tissue after selectively blocking group I mGluRs (1-Aminoindan-1,5-dicarboxylic acid (AIDA)) or group II mGluRs (LY 341495) during unilateral auditory nerve stimulation. Changes in [Ca(2+)]i of NM neurons were measured using fura-2 ratiometric calcium imaging and the tissue was subsequently processed for Y10B immunoreactivity (Y10B-ir), an antibody that recognizes a ribosomal epitope. The group I mGluR antagonist blocked the activity-dependent regulation of both [Ca(2+)]i and Y10B-ir, but the group II antagonist blocked only the activity-dependent regulation of Y10B-ir. That is, even when group II receptors were blocked, stimulation continued to maintain low [Ca(2+)]i, but it did not maintain Y10B-ir. These results suggest a dissociation in how calcium and ribosomes are regulated in NM neurons and that ribosomes can be regulated through a mechanism that is independent of calcium regulation.

  7. Platelet activating factors alters calcium homeostasis in cultured vascular endothelial cells

    SciTech Connect

    Brock, T.A.; Gimbrone, M.A. Jr.

    1986-06-01

    Platelet activating factor (1-O-alkyl-2-acetyl-sn-glycerol-3-phosphorylcholine; PAF), a potent in vivo mediator of allergic and inflammatory reactions, induced a rapid (onset less than 30 s), concentration-dependent (threshold approximately 10(-11) M, half-maximal approximately 10(-10) M, maximal approximately 10(-8)-10(-7) M) efflux of /sup 45/Ca/sup 2 +/ from preloaded cultured bovine aortic endothelial cells (BAEC). In contrast, deacetylated and other PAF analogues were essentially ineffective. PAF (10(-7) M) was also shown to increase cytosolic free calcium (49 +/- 5%) in suspensions of quin 2 (calcium-sensitive fluorescent dye)-loaded BAEC. PAF-stimulated /sup 45/Ca/sup 2 +/ efflux was not blocked by aspirin treatment (100 or 500 microM, 30 min). In the absence of external calcium, PAF was still highly effective in stimulating unidirectional /sup 45/Ca/sup 2 +/ efflux, thus suggesting that PAF mobilized a sequestered pool of intracellular calcium. CV-3988, a PAF antagonist, inhibited PAF-stimulated /sup 45/Ca/sup 2 +/ efflux in a dose-dependent manner. Pretreatment of BAEC with PAF (10(-8) M, 15 min), but not with other PAF analogues, resulted in a decrease in subsequent PAF-stimulated /sup 45/Ca/sup 2 +/ efflux, thus suggesting an agonist-specific desensitization. PAF also stimulated a 30% net decrease in the equilibrium /sup 45/Ca/sup 2 +/ content of BAEC within 1 min, which gradually recovered to prestimulus levels in 10-15 min. PAF-stimulated /sup 45/Ca/sup 2 +/ efflux was also observed in endothelial cells cultured from human umbilical vein and baboon cephalic vein but not from cultured human dermal fibroblasts or bovine aortic smooth muscle. These studies provide direct evidence for agonist- and cell-specific effects of PAF on vascular endothelium.

  8. Calcium Imaging of Neuronal Activity in Drosophila Can Identify Anticonvulsive Compounds.

    PubMed

    Streit, Anne K; Fan, Yuen Ngan; Masullo, Laura; Baines, Richard A

    2016-01-01

    Although there are now a number of antiepileptic drugs (AEDs) available, approximately one-third of epilepsy patients respond poorly to drug intervention. The reasons for this are complex, but are probably reflective of the increasing number of identified mutations that predispose individuals to this disease. Thus, there is a clear requirement for the development of novel treatments to address this unmet clinical need. The existence of gene mutations that mimic a seizure-like behaviour in the fruit fly, Drosophila melanogaster, offers the possibility to exploit the powerful genetics of this insect to identify novel cellular targets to facilitate design of more effective AEDs. In this study we use neuronal expression of GCaMP, a potent calcium reporter, to image neuronal activity using a non-invasive and rapid method. Expression in motoneurons in the isolated CNS of third instar larvae shows waves of calcium-activity that pass between segments of the ventral nerve cord. Time between calcium peaks, in the same neurons, between adjacent segments usually show a temporal separation of greater than 200 ms. Exposure to proconvulsants (picrotoxin or 4-aminopyridine) reduces separation to below 200 ms showing increased synchrony of activity across adjacent segments. Increased synchrony, characteristic of epilepsy, is similarly observed in genetic seizure mutants: bangsenseless1 (bss1) and paralyticK1270T (paraK1270T). Exposure of bss1 to clinically-used antiepileptic drugs (phenytoin or gabapentin) significantly reduces synchrony. In this study we use the measure of synchronicity to evaluate the effectiveness of known and novel anticonvulsive compounds (antipain, isethionate, etopiside rapamycin and dipyramidole) to reduce seizure-like CNS activity. We further show that such compounds also reduce the Drosophila voltage-gated persistent Na+ current (INaP) in an identified motoneuron (aCC). Our combined assays provide a rapid and reliable method to screen unknown compounds

  9. Calcium Imaging of Neuronal Activity in Drosophila Can Identify Anticonvulsive Compounds

    PubMed Central

    Streit, Anne K.; Fan, Yuen Ngan; Masullo, Laura; Baines, Richard A.

    2016-01-01

    Although there are now a number of antiepileptic drugs (AEDs) available, approximately one-third of epilepsy patients respond poorly to drug intervention. The reasons for this are complex, but are probably reflective of the increasing number of identified mutations that predispose individuals to this disease. Thus, there is a clear requirement for the development of novel treatments to address this unmet clinical need. The existence of gene mutations that mimic a seizure-like behaviour in the fruit fly, Drosophila melanogaster, offers the possibility to exploit the powerful genetics of this insect to identify novel cellular targets to facilitate design of more effective AEDs. In this study we use neuronal expression of GCaMP, a potent calcium reporter, to image neuronal activity using a non-invasive and rapid method. Expression in motoneurons in the isolated CNS of third instar larvae shows waves of calcium-activity that pass between segments of the ventral nerve cord. Time between calcium peaks, in the same neurons, between adjacent segments usually show a temporal separation of greater than 200 ms. Exposure to proconvulsants (picrotoxin or 4-aminopyridine) reduces separation to below 200 ms showing increased synchrony of activity across adjacent segments. Increased synchrony, characteristic of epilepsy, is similarly observed in genetic seizure mutants: bangsenseless1 (bss1) and paralyticK1270T (paraK1270T). Exposure of bss1 to clinically-used antiepileptic drugs (phenytoin or gabapentin) significantly reduces synchrony. In this study we use the measure of synchronicity to evaluate the effectiveness of known and novel anticonvulsive compounds (antipain, isethionate, etopiside rapamycin and dipyramidole) to reduce seizure-like CNS activity. We further show that such compounds also reduce the Drosophila voltage-gated persistent Na+ current (INaP) in an identified motoneuron (aCC). Our combined assays provide a rapid and reliable method to screen unknown compounds

  10. A highly sensitive fluorescent indicator dye for calcium imaging of neural activity in vitro and in vivo.

    PubMed

    Tada, Mayumi; Takeuchi, Atsuya; Hashizume, Miki; Kitamura, Kazuo; Kano, Masanobu

    2014-06-01

    Calcium imaging of individual neurons is widely used for monitoring their activity in vitro and in vivo. Synthetic fluorescent calcium indicator dyes are commonly used, but the resulting calcium signals sometimes suffer from a low signal-to-noise ratio (SNR). Therefore, it is difficult to detect signals caused by single action potentials (APs) particularly from neurons in vivo. Here we showed that a recently developed calcium indicator dye, Cal-520, is sufficiently sensitive to reliably detect single APs both in vitro and in vivo. In neocortical neurons, calcium signals were linearly correlated with the number of APs, and the SNR was > 6 for in vitro slice preparations and > 1.6 for in vivo anesthetised mice. In cerebellar Purkinje cells, dendritic calcium transients evoked by climbing fiber inputs were clearly observed in anesthetised mice with a high SNR and fast decay time. These characteristics of Cal-520 are a great advantage over those of Oregon Green BAPTA-1, the most commonly used calcium indicator dye, for monitoring the activity of individual neurons both in vitro and in vivo.

  11. Mitochondrial fusion dynamics is robust in the heart and depends on calcium oscillations and contractile activity.

    PubMed

    Eisner, Verónica; Cupo, Ryan R; Gao, Erhe; Csordás, György; Slovinsky, William S; Paillard, Melanie; Cheng, Lan; Ibetti, Jessica; Chen, S R Wayne; Chuprun, J Kurt; Hoek, Jan B; Koch, Walter J; Hajnóczky, György

    2017-01-31

    Mitochondrial fusion is thought to be important for supporting cardiac contractility, but is hardly detectable in cultured cardiomyocytes and is difficult to directly evaluate in the heart. We overcame this obstacle through in vivo adenoviral transduction with matrix-targeted photoactivatable GFP and confocal microscopy. Imaging in whole rat hearts indicated mitochondrial network formation and fusion activity in ventricular cardiomyocytes. Promptly after isolation, cardiomyocytes showed extensive mitochondrial connectivity and fusion, which decayed in culture (at 24-48 h). Fusion manifested both as rapid content mixing events between adjacent organelles and slower events between both neighboring and distant mitochondria. Loss of fusion in culture likely results from the decline in calcium oscillations/contractile activity and mitofusin 1 (Mfn1), because (i) verapamil suppressed both contraction and mitochondrial fusion, (ii) after spontaneous contraction or short-term field stimulation fusion activity increased in cardiomyocytes, and (iii) ryanodine receptor-2-mediated calcium oscillations increased fusion activity in HEK293 cells and complementing changes occurred in Mfn1. Weakened cardiac contractility in vivo in alcoholic animals is also associated with depressed mitochondrial fusion. Thus, attenuated mitochondrial fusion might contribute to the pathogenesis of cardiomyopathy.

  12. A large-conductance calcium-activated potassium channel in potato (Solanum tuberosum) tuber mitochondria.

    PubMed

    Koszela-Piotrowska, Izabela; Matkovic, Karolina; Szewczyk, Adam; Jarmuszkiewicz, Wieslawa

    2009-11-11

    In the present study, we describe the existence of a novel potassium channel in the plant [potato (Solanum tuberosum) tuber] mitochondrial inner membrane. We found that substances known to modulate large-conductance calcium-activated potassium channel activity influenced the bioenergetics of potato tuber mitochondria. In isolated mitochondria, Ca2+ and NS1619 {1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-ben-zimidazole-2-one; a potassium channel opener} were found to depolarize the mitochondrial membrane potential and to stimulate resting respiration. These effects were blocked by iberiotoxin (a potassium channel inhibitor) in a potassium-dependent manner. Additionally, the electrophysiological properties of the large-conductance potassium channel present in the potato tuber inner mitochondrial membrane are described in a reconstituted system, using planar lipid bilayers. After incorporation in 50/450 mM KCl gradient solutions, we recorded large-conductance potassium channel activity with conductance from 502+/-15 to 615+/-12 pS. The probability of channel opening was increased by Ca2+ and reduced by iberiotoxin. Immunological analysis with antibodies raised against the mammalian plasma-membrane large-conductance Ca2+-dependent K+ channel identified a pore-forming alpha subunit and an auxiliary beta2 subunit of the channel in potato tuber mitochondrial inner membrane. These results suggest that a large-conductance calcium-activated potassium channel similar to that of mammalian mitochondria is present in potato tuber mitochondria.

  13. Melanocortin 4 receptor constitutive activity inhibits L-type voltage-gated calcium channels in neurons.

    PubMed

    Agosti, F; Cordisco Gonzalez, S; Martinez Damonte, V; Tolosa, M J; Di Siervi, N; Schioth, H B; Davio, C; Perello, M; Raingo, J

    2017-03-27

    The melanocortin 4 receptor (MC4R) is a G protein-coupled receptor (GPCR) that is expressed in several brain nuclei playing a crucial role in the regulation of energy balance controlling the homeostasis of the organism. It displays both agonist-evoked and constitutive activity, and moreover, it can couple to different G proteins. Most of the research on MC4R has been focused on agonist-induced activity, while the molecular and cellular basis of MC4R constitutive activity remains scarcely studied. We have previously shown that neuronal N-type voltage-gated calcium channels (CaV2.2) are inhibited by MC4R agonist-dependent activation, while the CaV subtypes that carry L- and P/Q-type current are not. Here, we tested the hypothesis that MC4R constitutive activity can affect CaV, with focus on the channel subtypes that can control transcriptional activity coupled to depolarization (L-type, CaV1.2/1.3) and neurotransmitter release (N- and P/Q-type, CaV2.2 and CaV2.1). We found that MC4R constitutive activity inhibits specifically CaV1.2/1.3 and CaV2.1 subtypes of CaV. We also explored the signaling pathways mediating this inhibition, and thus propose that agonist-dependent and basal MC4R activation modes signal differentially through Gs and Gi/o pathways to impact on different CaV subtypes. In addition, we found that chronic incubation with MC4R endogenous inverse agonist, agouti and agouti-related peptide (AgRP), occludes CaV inhibition in a cell line and in amygdaloid complex cultured neurons as well. Thus, we define new mechanisms of control of the main mediators of depolarization-induced calcium entry into neurons by a GPCR that displays constitutive activity.

  14. Calcium regulation of HCN channels supports persistent activity in a multiscale model of neocortex

    PubMed Central

    McDougal, Robert A.; Bulanova, Anna S.; Zeki, Mustafa; Lakatos, Peter; Terman, David; Hines, Michael L.; Lytton, William W.

    2016-01-01

    Neuronal persistent activity has been primarily assessed in terms of electrical mechanisms, without attention to the complex array of molecular events that also control cell excitability. We developed a multiscale neocortical model proceeding from the molecular to the network level to assess the contributions of calcium regulation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in providing additional and complementary support of continuing activation in the network. The network contained 776 compartmental neurons arranged in the cortical layers, connected using synapses containing AMPA/NMDA/GABAA/GABAB receptors. Metabotropic glutamate receptors (mGluR) produced inositol triphosphate (IP3) which caused release of Ca2+ from endoplasmic reticulum (ER) stores, with reuptake by sarco/ER Ca2+-ATP-ase pumps (SERCA), and influence on HCN channels. Stimulus-induced depolarization led to Ca2+ influx via NMDA and voltage-gated Ca2+ channels (VGCCs). After a delay, mGluR activation led to ER Ca2+ release via IP3 receptors. These factors increased HCN channel conductance and produced firing lasting for ~1 minute. The model displayed inter-scale synergies among synaptic weights, excitation/inhibition balance, firing rates, membrane depolarization, calcium levels, regulation of HCN channels, and induction of persistent activity. The interaction between inhibition and Ca2+ at the HCN channel nexus determined a limited range of inhibition strengths for which intracellular Ca2+ could prepare population-specific persistent activity. Interactions between metabotropic and ionotropic inputs to the neuron demonstrated how multiple pathways could contribute in a complementary manner to persistent activity. Such redundancy and complementarity via multiple pathways is a critical feature of biological systems. Mediation of activation at different time scales, and through different pathways, would be expected to protect against disruption, in this case providing

  15. Duration differences of corticostriatal responses in striatal projection neurons depend on calcium activated potassium currents

    PubMed Central

    Arias-García, Mario A.; Tapia, Dagoberto; Flores-Barrera, Edén; Pérez-Ortega, Jesús E.; Bargas, José; Galarraga, Elvira

    2013-01-01

    The firing of striatal projection neurons (SPNs) exhibits afterhyperpolarizing potentials (AHPs) that determine discharge frequency. They are in part generated by Ca2+-activated K+-currents involving BK and SK components. It has previously been shown that suprathreshold corticostriatal responses are more prolonged and evoke more action potentials in direct pathway SPNs (dSPNs) than in indirect pathway SPNs (iSPNs). In contrast, iSPNs generate dendritic autoregenerative responses. Using whole cell recordings in brain slices, we asked whether the participation of Ca2+-activated K+-currents plays a role in these responses. Secondly, we asked if these currents may explain some differences in synaptic integration between dSPNs and iSPNs. Neurons obtained from BAC D1 and D2 GFP mice were recorded. We used charybdotoxin and apamin to block BK and SK channels, respectively. Both antagonists increased the depolarization and delayed the repolarization of suprathreshold corticostriatal responses in both neuron classes. We also used NS 1619 and NS 309 (CyPPA), to enhance BK and SK channels, respectively. Current enhancers hyperpolarized and accelerated the repolarization of corticostriatal responses in both neuron classes. Nevertheless, these drugs made evident that the contribution of Ca2+-activated K+-currents was different in dSPNs as compared to iSPNs: in dSPNs their activation was slower as though calcium took a diffusion delay to activate them. In contrast, their activation was fast and then sustained in iSPNs as though calcium flux activates them at the moment of entry. The blockade of Ca2+-activated K+-currents made iSPNs to look as dSPNs. Conversely, their enhancement made dSPNs to look as iSPNs. It is concluded that Ca2+-activated K+-currents are a main intrinsic determinant causing the differences in synaptic integration between corticostriatal polysynaptic responses between dSPNs and iSPNs. PMID:24109439

  16. Activation of phosphorylase by anoxia and dinitrophenol in rabbit colon smooth muscle: relation to release of calcium from mitochondria.

    PubMed

    Pettersson, G

    1983-05-01

    The effect of anoxia or 2,4-dinitrophenol (DNP) on the phosphorylase a activity and the calcium content in subcellular fractions from rabbit colon smooth muscle was studied. Anoxia for 15 min. as well as DNP (6.6 X 10(-5) M) for 5 min. increased the phosphorylase a activity. The calcium content in the mitochondrial subfraction, prepared from the anoxic- or DNP-treated intact muscle and determined by atomic absorption spectroscopy, was reduced. The calcium content in the nuclear and the microsomal fractions was not changed in preparations with a normal Ca-content. When the muscle was incubated for 60 min. in a Ca2+-free medium containing 2.0 mM EGTA, the calcium content in the mitochondrial fraction was reduced to 38% of the control. This calcium level was still further reduced and the phosphorylase a activity was increased by DNP in this "Ca-poor" muscle. In these preparations the Ca-content of the microsomal + supernatant fraction increased. Only when the muscle was incubated, initially, in an anoxic medium containing 0.1 mM Ca2+ for 120 min. and, subsequently, in an oxygenated medium containing 0.1 mM Ca2+ for 20 min., DNP failed to activate phosphorylase and to decrease the calcium content in the mitochondrial fraction. These results indicate that mitochondrial Ca2+ release is one of the regulatory factors of the anoxic-induced glycogenolysis.

  17. STIM1 calcium sensor is required for activation of the phagocyte oxidase during inflammation and host defense.

    PubMed

    Zhang, Hong; Clemens, Regina A; Liu, Fengchun; Hu, Yongmei; Baba, Yoshihiro; Theodore, Pierre; Kurosaki, Tomohiro; Lowell, Clifford A

    2014-04-03

    The stromal-interacting molecule 1 (STIM1) is a potent sensor of intracellular calcium, which in turn regulates entry of external calcium through plasma membrane channels to affect immune cell activation. Although the contribution of STIM1 to calcium signaling in lymphocytes has been well studied, the role of this protein in neutrophil-mediated inflammation and host defense is unknown. We report that STIM1-deficient murine neutrophils show loss of store-operated calcium entry (SOCE) in response to both soluble ligands that activate G-proteins as well as Fcγ-receptor or integrin ligation that activates tyrosine kinase signaling. This results in modest defects in phagocytosis and degranulation responses but a profound block in superoxide production by the phagocyte oxidase. We trace the primary intracellular target of calcium to be protein kinase C isoforms α and β (PKCα and PKCβ), which in turn phosphorylate subunits of the oxidase leading to superoxide production. In vivo the loss of SOCE in stim1(-/-) chimeric mice results in marked susceptibility to bacterial infections but also protection from tissue injury in hepatic ischemia/reperfusion injury. These results demonstrate the critical role of STIM1-mediated SOCE and define major protein targets of calcium signaling in neutrophil activation during inflammatory disease.

  18. Active overbank deposition during the last century, South River, Virginia

    NASA Astrophysics Data System (ADS)

    Pizzuto, Jim; Skalak, Katherine; Pearson, Adam; Benthem, Adam

    2016-03-01

    regional synthesis of floodplain deposition rates since European settlement. Floodplain sedimentation rates were high following European settlement, with published estimates ranging from 50 to 200 cm/100 years. Sedimentation rates decreased by 1 to 2 orders of magnitude during the twentieth and twenty-first centuries; but despite these lower sedimentation rates, floodplains continue to store a significant fraction of total suspended sediment load. Many floodplains of the mid-Atlantic region are active landforms fully connected to the rivers that flow within them and should not be considered terraces isolated from contemporary fluvial processes by post-settlement aggradation.

  19. Hypoxia-Inducible Factor-1α Causes Renal Cyst Expansion through Calcium-Activated Chloride Secretion

    PubMed Central

    Schley, Gunnar; Faria, Diana; Kroening, Sven; Willam, Carsten; Schreiber, Rainer; Klanke, Bernd; Burzlaff, Nicolai; Jantsch, Jonathan; Kunzelmann, Karl; Eckardt, Kai-Uwe

    2014-01-01

    Polycystic kidney diseases are characterized by numerous bilateral renal cysts that continuously enlarge and, through compression of intact nephrons, lead to a decline in kidney function over time. We previously showed that cyst enlargement is accompanied by regional hypoxia, which results in the stabilization of hypoxia-inducible transcription factor-1α (HIF-1α) in the cyst epithelium. Here we demonstrate a correlation between cyst size and the expression of the HIF-1α–target gene, glucose transporter 1, and report that HIF-1α promotes renal cyst growth in two in vitro cyst models—principal-like MDCK cells (plMDCKs) within a collagen matrix and cultured embryonic mouse kidneys stimulated with forskolin. In both models, augmenting HIF-1α levels with the prolyl hydroxylase inhibitor 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate enhanced cyst growth. In addition, inhibition of HIF-1α degradation through tubule-specific knockdown of the von Hippel-Lindau tumor suppressor increased cyst size in the embryonic kidney cyst model. In contrast, inhibition of HIF-1α by chetomin and knockdown of HIF-1α both decreased cyst growth in these models. Consistent with previous reports, plMDCK cyst enlargement was driven largely by transepithelial chloride secretion, which consists, in part, of a calcium-activated chloride conductance. plMDCKs deficient for HIF-1α almost completely lacked calcium-activated chloride secretion. We conclude that regional hypoxia in renal cysts contributes to cyst growth, primarily due to HIF-1α–dependent calcium-activated chloride secretion. These findings identify the HIF system as a novel target for inhibition of cyst growth. PMID:24203996

  20. Characterization of cytolytic neutrophil activation in vitro by amorphous hydrated calcium phosphate as a model of biomaterial inflammation.

    PubMed

    Edwards, Felicity C; Taheri, Amir; Dann, Sophie C; Dye, Julian F

    2011-03-01

    Calcium ions are utilized in biomolecular biomaterial design for osteomimetic scaffolds and as divalent cross-linking agents, typically for gelation of alginates, stabilisation of protein structure (e.g., fibrinogen) and enzyme activation (e.g., thrombin). Biological interactions with defined calcium phosphates (e.g., hydroxyapatite) are exploited for osteogenesis, although crystalline calcium phosphates (e.g., calcium pyrophosphate) stimulate inflammation. We found that the calcium concentration used in the manufacture of prototype dermal scaffolds made from fibrin/alginate composite was related to the inflammatory infiltration during in vivo integration. In investigating a cause for this inflammatory response, we have identified and characterized a cytolytic inflammatory effect of amorphous calcium phosphate (CaP) formed in physiological solutions, relevant to biomaterial biocompatibility. Isolated human neutrophils (Nφ) were incubated in phosphate-buffered saline with CaCl(2) ranging 2.5-20 mM total calcium. Nφ activation was assessed by morphology and integrin-β2 (CD18a) expression. Mediator release (Nφ-elastase, IL-8, and TNFα) was measured from both Nφ and whole blood cultures plus CaCl(2). CaP exposure increased CD18a expression over 1 h (maximal at 10 mM calcium/ phosphate) with concurrent phagocytosis, cytolysis, and Nφ-elastase release. CaCl(2) induced expression of IL-8 and TNFα in whole blood cultures. These results suggest that CaP formed from the resorption of calcium-containing biomaterials could induce inflammation and accelerate biomaterial degradation, driving further CaP release. This demonstrates a novel mechanism for biomaterial-induced inflammation. The in vitro system described could aid preclinical evaluation of novel biomaterial inflammatory potential.

  1. Contribution of presynaptic calcium-activated potassium currents to transmitter release regulation in cultured Xenopus nerve-muscle synapses.

    PubMed

    Pattillo, J M; Yazejian, B; DiGregorio, D A; Vergara, J L; Grinnell, A D; Meriney, S D

    2001-01-01

    Using Xenopus nerve-muscle co-cultures, we have examined the contribution of calcium-activated potassium (K(Ca)) channels to the regulation of transmitter release evoked by single action potentials. The presynaptic varicosities that form on muscle cells in these cultures were studied directly using patch-clamp recording techniques. In these developing synapses, blockade of K(Ca) channels with iberiotoxin or charybdotoxin decreased transmitter release by an average of 35%. This effect would be expected to be caused by changes in the late phases of action potential repolarization. We hypothesize that these changes are due to a reduction in the driving force for calcium that is normally enhanced by the local hyperpolarization at the active zone caused by potassium current through the K(Ca) channels that co-localize with calcium channels. In support of this hypothesis, we have shown that when action potential waveforms were used as voltage-clamp commands to elicit calcium current in varicosities, peak calcium current was reduced only when these waveforms were broadened beginning when action potential repolarization was 20% complete. In contrast to peak calcium current, total calcium influx was consistently increased following action potential broadening. A model, based on previously reported properties of ion channels, faithfully reproduced predicted effects on action potential repolarization and calcium currents. From these data, we suggest that the large-conductance K(Ca) channels expressed at presynaptic varicosities regulate transmitter release magnitude during single action potentials by altering the rate of action potential repolarization, and thus the magnitude of peak calcium current.

  2. International Union of Basic and Clinical Pharmacology. LXXXV: Calcium-Activated Chloride Channels

    PubMed Central

    Huang, Fen; Wong, Xiuming

    2012-01-01

    Calcium-activated chloride channels (CaCCs) are widely expressed in various tissues and implicated in physiological processes such as sensory transduction, epithelial secretion, and smooth muscle contraction. Transmembrane proteins with unknown function 16 (TMEM16A) has recently been identified as a major component of CaCCs. Detailed molecular analysis of TMEM16A will be needed to understand its structure-function relationships. The role this channel plays in physiological systems remains to be established and is currently a subject of intense investigation. PMID:22090471

  3. Calcium phosphate deposition rate, structure and osteoconductivity on electrospun poly(l-lactic acid) matrix using electrodeposition or simulated body fluid incubation.

    PubMed

    He, Chuanglong; Jin, Xiaobing; Ma, Peter X

    2014-01-01

    Mineralized nanofibrous scaffolds have been proposed as promising scaffolds for bone regeneration due to their ability to mimic both nanoscale architecture and chemical composition of natural bone extracellular matrix. In this study, a novel electrodeposition method was compared with an extensively explored simulated body fluid (SBF) incubation method in terms of the deposition rate, chemical composition and morphology of calcium phosphate formed on electrospun fibrous thin matrices with a fiber diameter in the range ~200-1400 nm prepared using 6, 8, 10 and 12 wt.% poly(l-lactic acid) (PLLA) solutions in a mixture of dichloromethane and acetone (2:1 in volume). The effects of the surface modification using the two mineralization techniques on osteoblastic cell (MC3T3-E1) proliferation and differentiation were also examined. It was found that electrodeposition was two to three orders of magnitude faster than the SBF method in mineralizing the fibrous matrices, reducing the mineralization time from ~2 weeks to 1h to achieve the same amounts of mineralization. The mineralization rate also varied with the fiber diameter but in opposite directions between the two mineralization methods. As a general trend, the increase of fiber diameter resulted in a faster mineralization rate for the electrodeposition method but a slower mineralization rate for the SBF incubation method. Using the electrodeposition method, one can control the chemical composition and morphology of the calcium phosphate by varying the electric deposition potential and electrolyte temperature to tune the mixture of dicalcium phosphate dihydrate and hydroxyapatite (HAp). Using the SBF method, one can only obtain a low crystallinity HAp. The mineralized electrospun PLLA fibrous matrices from either method similarly facilitate the proliferation and osteogenic differentiation of preosteoblastic MC3T3-E1 cells as compared to neat PLLA matrices. Therefore, the electrodeposition method can be utilized as a fast

  4. Osteoclast spreading kinetics are correlated with an oscillatory activation of a calcium-dependent potassium current.

    PubMed

    Espinosa, Leon; Paret, Laurent; Ojeda, Carlos; Tourneur, Yves; Delmas, Pierre D; Chenu, Chantal

    2002-10-01

    Cell movement and spreading involve calcium-dependent processes and ionic channel activation. During bone resorption, osteoclasts alternate between spread, motile and resorptive phases. We investigated whether the electrical membrane properties of osteoclasts were linked to their membrane morphological changes. Rabbit osteoclasts were recorded by time-lapse videomicroscopy performed simultaneously with patch-clamp whole cell and single channel recordings. Original image analysis methods were developed and used to demonstrate for the first time an oscillatory activation of a spontaneous membrane current in osteoclasts, which is directly correlated to the membrane movement rate. This current was identified as a calcium-dependent potassium current (IK(Ca)) that is sensitive to both charybdotoxin and apamin and was generated by a channel with unitary conductance of approximately 25+/-2 pS. Blockade of this current also decreased osteoclast spreading and inhibited bone resorption in vitro, demonstrating a physiological role for this current in osteoclast activity. These results establish for the first time a temporal correlation between lamellipodia formation kinetics and spontaneous peaks of IK(Ca), which are both involved in the control of osteoclast spreading and bone resorption.

  5. EFFECTS OF SODIUM AND CALCIUM IN LIGNITE ON THE PERFORMANCE OF ACTIVATED CARBON PRODUCTS

    SciTech Connect

    Edwin S. Olson; Kurt E. Eylands; Daniel J. Stepan

    2001-12-01

    Powdered activated carbon (PAC) has traditionally been used by the water treatment industry for the removal of compounds contributing to taste and odor problems. PAC also has the potential to remove naturally occurring organic matter (NOM) from raw waters prior to disinfection, thus controlling the formation of regulated disinfection by-products (DBPs). Many small water systems are currently using PAC for taste and odor control and have the potential to use PAC for controlling DBPs. The Energy & Environmental Research Center has been working on the development of a PAC product to remove NOM from surface water supplies to prevent the formation of carcinogenic DBPs during chlorination. During previous studies, the sodium and calcium content of the lignites showed a significant effect on the sorption capacity of the activated carbon product. As much as a 130% increase in the humic acid sorption capacity of a PAC produced from a high-sodium-content lignite was observed. During this study, activated carbons were prepared from three coals representing high-sodium, low-sodium--low-calcium, and high-calcium compositions in two steps, an initial char formation followed by mild activation with steam to avoid excessive burnout. This set of carbons was characterized with respect to physical and chemical properties. The BET (Brunauer-Emmett-Teller) nitrogen adsorption isotherms gave relatively low surface areas (ranging from 245 to 370 m{sup 2}/g). The lowest-BET area was obtained for the high-sodium carbon, which can be attributed to enlargement of micropores as a result of sodium-catalyzed gasification reaction of the carbon structure. This hypothesis is consistent with the scanning electron microscopy microprobe analyses, which show that in both the coal and the activated carbon from this coal, the sodium is distributed over both the carbon structure and the mineral particles. Thus it is initially associated with carboxylate groups on the coal and then as sodium oxide or

  6. Effects of calcium infusion on secretion and motor activity of totally isolated canine stomach perfused with homologous blood.

    PubMed

    Kowalewski, K; Kolodej, A

    1976-01-01

    Isolated, ex vivo perfused, canine stomachs were used for this study. Gastric secretion, myoelectrical activity and mechanical activity were recorded during stimulation of gastric function with pentagastrin or histamine alone or combined with calcium gluconate. Secretagogues and calcium were infused into the gastric arterial circulation. Hypercalcemia induced significant inhibition of pentagastrin, stimulated gastric secretion, but did not affect the secretion stimulated by histamine. Hypercalcemia also induced an increase of frequency of cycles of electrical control activity and a decrease of mechanical activity of the gastric antrum. The effect of hypercalcemia on gastric motor function was similar in the nonstimulated stomach and during the infusion of secretagogues used in this experiment.

  7. Genetically encoded calcium indicators for multi-color neural activity imaging and combination with optogenetics

    PubMed Central

    Akerboom, Jasper; Carreras Calderón, Nicole; Tian, Lin; Wabnig, Sebastian; Prigge, Matthias; Tolö, Johan; Gordus, Andrew; Orger, Michael B.; Severi, Kristen E.; Macklin, John J.; Patel, Ronak; Pulver, Stefan R.; Wardill, Trevor J.; Fischer, Elisabeth; Schüler, Christina; Chen, Tsai-Wen; Sarkisyan, Karen S.; Marvin, Jonathan S.; Bargmann, Cornelia I.; Kim, Douglas S.; Kügler, Sebastian; Lagnado, Leon; Hegemann, Peter; Gottschalk, Alexander; Schreiter, Eric R.; Looger, Loren L.

    2013-01-01

    Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, “RCaMPs,” engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca2+-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca2+]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca2+ affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics. PMID:23459413

  8. Neuroprotective activity of stiripentol with a possible involvement of voltage-dependent calcium and sodium channels.

    PubMed

    Verleye, Marc; Buttigieg, Dorothée; Steinschneider, Rémy

    2016-02-01

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

  9. Reporting sodium channel activity using calcium flux: pharmacological promiscuity of cardiac Nav1.5.

    PubMed

    Zhang, Hongkang; Zou, Beiyan; Du, Fang; Xu, Kaiping; Li, Min

    2015-02-01

    Voltage-gated sodium (Nav) channels are essential for membrane excitability and represent therapeutic targets for treating human diseases. Recent reports suggest that these channels, e.g., Nav1.3 and Nav1.5, are inhibited by multiple structurally distinctive small molecule drugs. These studies give reason to wonder whether these drugs collectively target a single site or multiple sites in manifesting such pharmacological promiscuity. We thus investigate the pharmacological profile of Nav1.5 through systemic analysis of its sensitivity to diverse compound collections. Here, we report a dual-color fluorescent method that exploits a customized Nav1.5 [calcium permeable Nav channel, subtype 5 (SoCal5)] with engineered-enhanced calcium permeability. SoCal5 retains wild-type (WT) Nav1.5 pharmacological profiles. WT SoCal5 and SoCal5 with the local anesthetics binding site mutated (F1760A) could be expressed in separate cells, each with a different-colored genetically encoded calcium sensor, which allows a simultaneous report of compound activity and site dependence. The pharmacological profile of SoCal5 reveals a hit rate (>50% inhibition) of around 13% at 10 μM, comparable to that of hERG. The channel activity is susceptible to blockage by known drugs and structurally diverse compounds. The broad inhibition profile is highly dependent on the F1760 residue in the inner cavity, which is a residue conserved among all nine subtypes of Nav channels. Both promiscuity and dependence on F1760 seen in Nav1.5 were replicated in Nav1.4. Our evidence of a broad inhibition profile of Nav channels suggests a need to consider off-target effects on Nav channels. The site-dependent promiscuity forms a foundation to better understand Nav channels and compound interactions.

  10. Functional segregation of voltage-activated calcium channels in motoneurons of the dorsal motor nucleus of the vagus

    PubMed Central

    Cooper, Garry; Lasser-Katz, Efrat; Simchovitz, Alon; Sharon, Ronit; Soreq, Hermona; Surmeier, D. James

    2015-01-01

    Calcium influx elevates mitochondrial oxidant stress (mOS) in dorsal motor nucleus of the vagus (DMV) neurons that are prone to Lewy body pathologies in presymptomatic Parkinson's disease (PD) patients. In experimental PD models, treatment with isradipine, the dihydropyridine with the highest affinity to Cav1.3 channels, prevents subthreshold calcium influx via Cav1.3 channels into midbrain dopamine neurons and protects them from mOS. In DMV neurons, isradipine is also effective in reducing mOS despite overwhelming evidence that subthreshold calcium influx is negligible compared with spike-triggered influx. To solve this conundrum we combined slice electrophysiology, two-photon laser scanning microscopy, mRNA profiling, and computational modeling. We find that the unusually depolarized subthreshold voltage trajectory of DMV neurons is positioned between the relatively hyperpolarized activation curve of Cav1.3 channels and that of other high-voltage activated (HVA) calcium channels, thus creating a functional segregation between Cav1.3 and HVA calcium channels. The HVA channels flux the bulk of calcium during spikes but can only influence pacemaking through their coupling to calcium-activated potassium currents. In contrast, Cav1.3 currents, which we show to be more than an order-of-magnitude smaller than the HVA calcium currents, are able to introduce sufficient inward current to speed up firing. However, Kv4 channels that are constitutively open in the subthreshold range guarantee slow pacemaking, despite the depolarizing action of Cav1.3 and other pacemaking currents. We propose that the efficacy of isradipine in preventing mOS in DMV neurons arises from its mixed effect on Cav1.3 channels and on HVA Cav1.2 channels. PMID:26156385

  11. Functional segregation of voltage-activated calcium channels in motoneurons of the dorsal motor nucleus of the vagus.

    PubMed

    Cooper, Garry; Lasser-Katz, Efrat; Simchovitz, Alon; Sharon, Ronit; Soreq, Hermona; Surmeier, D James; Goldberg, Joshua A

    2015-09-01

    Calcium influx elevates mitochondrial oxidant stress (mOS) in dorsal motor nucleus of the vagus (DMV) neurons that are prone to Lewy body pathologies in presymptomatic Parkinson's disease (PD) patients. In experimental PD models, treatment with isradipine, the dihydropyridine with the highest affinity to Cav1.3 channels, prevents subthreshold calcium influx via Cav1.3 channels into midbrain dopamine neurons and protects them from mOS. In DMV neurons, isradipine is also effective in reducing mOS despite overwhelming evidence that subthreshold calcium influx is negligible compared with spike-triggered influx. To solve this conundrum we combined slice electrophysiology, two-photon laser scanning microscopy, mRNA profiling, and computational modeling. We find that the unusually depolarized subthreshold voltage trajectory of DMV neurons is positioned between the relatively hyperpolarized activation curve of Cav1.3 channels and that of other high-voltage activated (HVA) calcium channels, thus creating a functional segregation between Cav1.3 and HVA calcium channels. The HVA channels flux the bulk of calcium during spikes but can only influence pacemaking through their coupling to calcium-activated potassium currents. In contrast, Cav1.3 currents, which we show to be more than an order-of-magnitude smaller than the HVA calcium currents, are able to introduce sufficient inward current to speed up firing. However, Kv4 channels that are constitutively open in the subthreshold range guarantee slow pacemaking, despite the depolarizing action of Cav1.3 and other pacemaking currents. We propose that the efficacy of isradipine in preventing mOS in DMV neurons arises from its mixed effect on Cav1.3 channels and on HVA Cav1.2 channels.

  12. Low-temperature SiON films deposited by plasma-enhanced atomic layer deposition method using activated silicon precursor

    SciTech Connect

    Suh, Sungin; Kim, Jun-Rae; Kim, Seongkyung; Hwang, Cheol Seong; Kim, Hyeong Joon; Ryu, Seung Wook; Cho, Seongjae

    2016-01-15

    It has not been an easy task to deposit SiN at low temperature by conventional plasma-enhanced atomic layer deposition (PE-ALD) since Si organic precursors generally have high activation energy for adsorption of the Si atoms on the Si-N networks. In this work, in order to achieve successful deposition of SiN film at low temperature, the plasma processing steps in the PE-ALD have been modified for easier activation of Si precursors. In this modification, the efficiency of chemisorption of Si precursor has been improved by additional plasma steps after purging of the Si precursor. As the result, the SiN films prepared by the modified PE-ALD processes demonstrated higher purity of Si and N atoms with unwanted impurities such as C and O having below 10 at. % and Si-rich films could be formed consequently. Also, a very high step coverage ratio of 97% was obtained. Furthermore, the process-optimized SiN film showed a permissible charge-trapping capability with a wide memory window of 3.1 V when a capacitor structure was fabricated and measured with an insertion of the SiN film as the charge-trap layer. The modified PE-ALD process using the activated Si precursor would be one of the most practical and promising solutions for SiN deposition with lower thermal budget and higher cost-effectiveness.

  13. Imaging spinal neuron ensembles active during locomotion with genetically encoded calcium indicators.

    PubMed

    Hinckley, Christopher A; Pfaff, Samuel L

    2013-03-01

    Advances in molecular-genetic tools for labeling neuronal subtypes, and the emerging development of robust genetic probes for neural activity, are likely to revolutionize our understanding of the functional organization of neural circuits. In principle, these tools should be able to detect activity at cellular resolution for large ensembles of identified neuron types as they participate in specific behaviors. This report describes the use of genetically encoded calcium indicators (GECIs), combined with two-photon microscopy, to characterize V1 interneurons, known to be critical for setting the duration of the step cycle. All V1 interneurons arise from a common precursor population and express engrailed-1 (En1). Our data show that although neighboring interneurons that arise from the same developmental lineage and share many features, such as projection patterns and neurotransmitter profiles, they are not irrevocably committed to having the same pattern of activity.

  14. Inertisation of galvanic sludge with calcium oxide, activated carbon, and phosphoric acid.

    PubMed

    Oreščanin, Višnja; Lovrenčić Mikelić, Ivanka; Kollar, Robert; Mikulić, Nenad; Medunić, Gordana

    2012-09-01

    In this study we compared three methods for the treatment of electroplating sludge highly loaded with zinc and iron: (1) calcium oxide-based solidification/stabilisation; (2) conversion into inert material by adsorption of organic and inorganic pollutants onto activated carbon; and (3) conversion of mobile waste components into insoluble phosphates. All three methods proved highly efficient in the conversion of hazardous waste into inert material. Under optimum treatment conditions zinc concentration in the leachate of solidified waste was reduced by 99.7 % compared to untreated sludge. Zinc retention efficiency in the waste treated with activated carbon and phosphoric acid was 99.9 % and 98.7 %, respectively. The advantages of electroplating sludge treatment with activated carbon over the other two methods are high sorption capacity, insignificant pH and volume changes of the sludge, and simple use.

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

    PubMed

    Kaur, Mandeep; Goel, Rajesh Kumar

    2011-01-01

    "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.

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

    PubMed Central

    Kaur, Mandeep; Goel, Rajesh Kumar

    2011-01-01

    “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. PMID:19948752

  17. Activation of Src and release of intracellular calcium by phosphatidic acid during Xenopus laevis fertilization.

    PubMed

    Bates, Ryan C; Fees, Colby P; Holland, William L; Winger, Courtney C; Batbayar, Khulan; Ancar, Rachel; Bergren, Todd; Petcoff, Douglas; Stith, Bradley J

    2014-02-01

    We report a new step in the fertilization in Xenopus laevis which has been found to involve activation of Src tyrosine kinase to stimulate phospholipase C-γ (PLC-γ) which increases inositol 1,4,5-trisphosphate (IP3) to release intracellular calcium ([Ca](i)). Molecular species analysis and mass measurements suggested that sperm activate phospholipase D (PLD) to elevate phosphatidic acid (PA). We now report that PA mass increased 2.7 fold by 1 min after insemination and inhibition of PA production by two methods inhibited activation of Src and PLCγ, increased [Ca](i) and other fertilization events. As compared to 14 other lipids, PA specifically bound Xenopus Src but not PLCγ. Addition of synthetic PA activated egg Src (an action requiring intact lipid rafts) and PLCγ as well as doubling the amount of PLCγ in rafts. In the absence of elevated [Ca](i), PA addition elevated IP3 mass to levels equivalent to that induced by sperm (but twice that achieved by calcium ionophore). Finally, PA induced [Ca](i) release that was blocked by an IP3 receptor inhibitor. As only PLD1b message was detected, and Western blotting did not detect PLD2, we suggest that sperm activate PLD1b to elevate PA which then binds to and activates Src leading to PLCγ stimulation, IP3 elevation and [Ca](i) release. Due to these and other studies, PA may also play a role in membrane fusion events such as sperm-egg fusion, cortical granule exocytosis, the elevation of phosphatidylinositol 4,5-bisphosphate and the large, late increase in sn 1,2-diacylglycerol in fertilization.

  18. Legal Deposit and Preservation of Digital Publications: A Review of Research and Development Activity.

    ERIC Educational Resources Information Center

    Muir, Adrienne

    2001-01-01

    This review of research and development work deals with activities specifically related to digital legal deposit. An overview of research activity in the various stages of the deposit process-particularly in digital preservation-follows. Building the infrastructure and pilot depositories are discussed. Issues that are not currently being addressed…

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

    PubMed Central

    Gómez, Melissa; González, Alberto; Sáez, Claudio A.; Morales, Bernardo; Moenne, Alejandra

    2015-01-01

    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 11 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 activation of CaMs and CDPKs. 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 that copper entry through TRP channels leads to membrane depolarization. Moreover, membrane depolarization events were inhibited by W-7 and staurosporine, indicating that activation of CaMs and CDPKs is required to allow copper entry through TRPs. Interestingly, copper-induced calcium increases and depolarization events were light-dependent and were inhibited by DCMU, an inhibitor of photosystem II, and ATP-γ-S, a non-hydrolizable analog of ATP, suggesting that ATP derived from photosynthesis is required to activate TRPs. Thus, light-dependent copper-induced activation TRPC5, A1

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

    PubMed

    Gómez, Melissa; González, Alberto; Sáez, Claudio A; Morales, Bernardo; Moenne, Alejandra

    2015-01-01

    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 11 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 activation of CaMs and CDPKs. 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 that copper entry through TRP channels leads to membrane depolarization. Moreover, membrane depolarization events were inhibited by W-7 and staurosporine, indicating that activation of CaMs and CDPKs is required to allow copper entry through TRPs. Interestingly, copper-induced calcium increases and depolarization events were light-dependent and were inhibited by DCMU, an inhibitor of photosystem II, and ATP-γ-S, a non-hydrolizable analog of ATP, suggesting that ATP derived from photosynthesis is required to activate TRPs. Thus, light-dependent copper-induced activation TRPC5, A1

  1. p21-activated kinase regulates mast cell degranulation via effects on calcium mobilization and cytoskeletal dynamics

    PubMed Central

    Allen, Jayme D.; Jaffer, Zahara M.; Park, Su-Jung; Burgin, Sarah; Hofmann, Clemens; Sells, Mary Ann; Chen, Shi; Derr-Yellin, Ethel; Michels, Elizabeth G.; McDaniel, Andrew; Bessler, Waylan K.; Ingram, David A.; Atkinson, Simon J.; Travers, Jeffrey B.

    2009-01-01

    Mast cells are key participants in allergic diseases via activation of high-affinity IgE receptors (FcϵRI) resulting in release of proinflammatory mediators. The biochemical pathways linking IgE activation to calcium influx and cytoskeletal changes required for intracellular granule release are incompletely understood. We demonstrate, genetically, that Pak1 is required for this process. In a passive cutaneous anaphylaxis experiment, Wsh/Wsh mast cell–deficient mice locally reconstituted with Pak1−/− bone marrow–derived mast cells (BMMCs) experienced strikingly decreased allergen-induced vascular permeability compared with controls. Consistent with the in vivo phenotype, Pak1−/− BMMCs exhibited a reduction in FcϵRI-induced degranulation. Further, Pak1−/− BMMCs demonstrated diminished calcium mobilization and altered depolymerization of cortical filamentous actin (F-actin) in response to FcϵRI stimulation. These data implicate Pak1 as an essential molecular target for modulating acute mast cell responses that contribute to allergic diseases. PMID:19124833

  2. A novel orange phosphor of Eu 2+-activated calcium chlorosilicate for white light-emitting diodes

    NASA Astrophysics Data System (ADS)

    Ding, Weijia; Wang, Jing; Zhang, Mei; Zhang, Qiuhong; Su, Qiang

    2006-11-01

    Novel orange phosphor of Eu 2+-activated calcium chlorosilicate was synthesized at 1273 K by conventional solid-state reactions under reductive atmosphere and investigated by means of photoluminescence excitation, diffuse reflectance and emission spectroscopies. These results show that this phosphor can be efficiently excited by the incident light of 300-450 nm, well matched with the emission band of 395 nm-emitting InGaN chip, and emits an intense orange light peaking at 585 nm. By combining this phosphor with a 395 nm-emitting InGaN chip, an intense orange light-emitting diode (LED) was fabricated. Under 20 mA forward-bias current, its CIE chromaticity coordinates are (0.486, 0.446). The dependence of as-fabricated orange LED on forward-bias current indicates that it shows excellent chromaticity stability and luminance saturation. These results show that this Eu 2+-activated calcium chlorosilicate is a promising orange-emitting phosphor for near-ultraviolet (UV) InGaN-based white LED.

  3. Dietary calcium restriction affects mesenchymal stem cell activity and bone development in neonatal pigs.

    PubMed

    Mahajan, Avanika; Alexander, Lindsey S; Seabolt, Brynn S; Catrambone, Daniel E; McClung, James P; Odle, Jack; Pfeiler, T Wayne; Loboa, Elizabeth G; Stahl, Chad H

    2011-03-01

    The effects of dietary calcium (Ca) deficiency on skeletal integrity are well characterized in growing and mature mammals; however, less is known about Ca nutrition during the neonatal period. In this study, we examined the effects of neonatal Ca nutrition on bone integrity, endocrine hormones, and mesenchymal stem cell (MSC) activity. Neonatal pigs (24 ± 6 h of age) received either a Ca-adequate (1.2 g/100 g) or an ~40% Ca-deficient diet for 18 d. Ca deficiency reduced (P < 0.05) bone flexural strength and bone mineral density without major differences in plasma indicators of Ca status. There were no meaningful differences in plasma Ca, phosphate (PO(4)), parathyroid hormone, or 1,25-dihydroxycholecalciferol due to Ca nutrition throughout the study. Calcium deficiency also reduced (P < 0.05) the in vivo proliferation of MSC by ~50%. In vitro studies utilizing homologous sera demonstrated that MSC activity was affected (P < 0.05) by both the Ca status of the pig and the sera as well as by their interaction. The results indicate that neonatal Ca nutrition is crucial for bone integrity and suggest that early-life Ca restriction may have long-term effects on bone integrity via programming of MSC.

  4. Adsorption of arsenic by activated carbon, calcium alginate and their composite beads.

    PubMed

    Hassan, A F; Abdel-Mohsen, A M; Elhadidy, H

    2014-07-01

    The present investigation deals with preparation of three different adsorbent materials namely; potassium hydroxide activated carbon based apricot stone (C), calcium alginate beads (G) and calcium alginate/activated carbon composite beads (GC) were used for the removal of arsenic. The prepared adsorbent materials were characterized by scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR), N2-adsorption at -196°C and point of zero charge. From the obtained results, it was found that the porosity, surface area and total pore volume of the adsorbent material C>GC>G respectively, however, the G adsorbent has more acidic function group than the other adsorbents. The influence of pH, time, temperature and initial concentration of arsenic(V) were studied and optimized. GC exhibits the maximum As(V) adsorption (66.7mg/g at 30°C). The adsorption of arsenic ions was observed to follow pseudo-second order mechanism as well as the thermodynamic parameters confirm also the endothermic spontaneous and a physisorption process.

  5. Population calcium imaging of spontaneous respiratory and novel motor activity in the facial nucleus and ventral brainstem in newborn mice

    PubMed Central

    Persson, Karin; Rekling, Jens C

    2011-01-01

    Abstract The brainstem contains rhythm and pattern forming circuits, which drive cranial and spinal motor pools to produce respiratory and other motor patterns. Here we used calcium imaging combined with nerve recordings in newborn mice to reveal spontaneous population activity in the ventral brainstem and in the facial nucleus. In Fluo-8 AM loaded brainstem–spinal cord preparations, respiratory activity on cervical nerves was synchronized with calcium signals at the ventrolateral brainstem surface. Individual ventrolateral neurons at the level of the parafacial respiratory group showed perfect or partial synchrony with respiratory nerve bursts. In brainstem–spinal cord preparations, cut at the level of the mid-facial nucleus, calcium signals were recorded in the dorsal, lateral and medial facial subnuclei during respiratory activity. Strong activity initiated in the dorsal subnucleus, followed by activity in lateral and medial subnuclei. Whole-cell recordings from facial motoneurons showed weak respiratory drives, and electrical field potential recordings confirmed respiratory drive to particularly the dorsal and lateral subnuclei. Putative facial premotoneurons showed respiratory-related calcium signals, and were predominantly located dorsomedial to the facial nucleus. A novel motor activity on facial, cervical and thoracic nerves was synchronized with calcium signals at the ventromedial brainstem extending from the level of the facial nucleus to the medulla–spinal cord border. Cervical dorsal root stimulation induced similar ventromedial activity. The medial facial subnucleus showed calcium signals synchronized with this novel motor activity on cervical nerves, and cervical dorsal root stimulation induced similar medial facial subnucleus activity. In conclusion, the dorsal and lateral facial subnuclei are strongly respiratory-modulated, and the brainstem contains a novel pattern forming circuit that drives the medial facial subnucleus and cervical motor

  6. Calcium supplements

    MedlinePlus

    ... TYPES OF CALCIUM SUPPLEMENTS Forms of calcium include: Calcium carbonate: Over-the-counter (OTC) antacid products, such as Tums and Rolaids, contain calcium carbonate. These sources of calcium do not cost much. ...

  7. [Soil enzyme activities under two forest types as affected by different levels of nitrogen deposition].

    PubMed

    Zhao, Yu-tao; Li, Xue-feng; Han, Shi-jie; Hu, Yan-ling

    2008-12-01

    A simulation test was conducted to study the change trends of soil cellulase, polyphenol oxidase, and sucrase activities under natural broadleaf-Korean pine (Pinus koraiensis) and secondary poplar (Populus davidiana) -birch (Betula platyphylla) mixed forests as affected by 0, 25, and 50 kg x hm(-2) x a(-1) of N deposition. The results showed that the effects of elevated N deposition on test enzyme activities varied with forest type, and short-term nitrogen addition could significantly affect the test enzyme activities. High N deposition decreased soil polyphyneol oxidase activity, and correspondingly, soil cellulase and sucrase activities also had a trend of decrease.

  8. Dual functions for the endoplasmic reticulum calcium sensors STIM1 and STIM2 in T cell activation and tolerance

    PubMed Central

    Oh-hora, Masatsugu; Yamashita, Megumi; Hogan, Patrick G; Sharma, Sonia; Lamperti, Ed; Chung, Woo; Prakriya, Murali; Feske, Stefan; Rao, Anjana

    2009-01-01

    Store-operated Ca2+ entry through calcium release–activated calcium channels is the chief mechanism for increasing intracellular Ca2+ in immune cells. Here we show that mouse T cells and fibroblasts lacking the calcium sensor STIM1 had severely impaired store-operated Ca2+ influx, whereas deficiency in the calcium sensor STIM2 had a smaller effect. However, T cells lacking either STIM1 or STIM2 had much less cytokine production and nuclear translocation of the transcription factor NFAT. T cell–specific ablation of both STIM1 and STIM2 resulted in a notable lymphoproliferative phenotype and a selective decrease in regulatory T cell numbers. We conclude that both STIM1 and STIM2 promote store-operated Ca2+ entry into T cells and fibroblasts and that STIM proteins are required for the development and function of regulatory T cells. PMID:18327260

  9. Cannabinoid Receptor Activation Modifies NMDA Receptor Mediated Release of Intracellular Calcium: Implications for Endocannabinoid Control of Hippocampal Neural Plasticity

    PubMed Central

    Hampson, Robert E.; Miller, Frances; Palchik, Guillermo; Deadwyler, Sam A.

    2011-01-01

    Chronic activation or inhibition of cannabinoid receptors (CB1) leads to continuous suppression of neuronal plasticity in hippocampus and other brain regions, suggesting that endocannabinoids may have a functional role in synaptic processes that produce state-dependent transient modulation of hippocampal cell activity. In support of this, it has previously been shown in vitro that cannabinoid CB1 receptors modulate second messenger systems in hippocampal neurons that can modulate intracellular ion channels, including channels which release calcium from intracellular stores. Here we demonstrate in hippocampal slices a similar endocannabinoid action on excitatory glutamatergic synapses via modulation of NMDA-receptor mediated intracellular calcium levels in confocal imaged neurons. Calcium entry through glutamatergic NMDA-mediated ion channels increases intracellular calcium concentrations via modulation of release from ryanodine-sensitive channels in endoplasmic reticulum. The studies reported here show that NMDA-elicited increases in Calcium Green fluorescence are enhanced by CB1 receptor antagonists (i.e. rimonabant), and inhibited by CB1 agonists (i.e. WIN 55,212-2). Suppression of endocannabinoid breakdown by either reuptake inhibition (AM404) or fatty-acid amide hydrolase inhibition (URB597) produced suppression of NMDA elicited calcium increases comparable to WIN 55,212-2, while enhancement of calcium release provoked by endocannabinoid receptor antagonists (Rimonabant) was shown to depend on the blockade of CB1 receptor mediated de-phosphorylation of Ryanodine receptors. Such CB1 receptor modulation of NMDA elicited increases in intracellular calcium may account for the respective disruption and enhancement by CB1 agents of trial-specific hippocampal neuron ensemble firing patterns during performance of a short-term memory task, reported previously from this laboratory. PMID:21288475

  10. Vitamin D and Calcium Intakes, Physical Activity, and Calcaneus BMC among School-Going 13-Year Old Malaysian Adolescents

    PubMed Central

    Suriawati, A. A.; Abdul Majid, Hazreen; Al-Sadat, Nabilla; Mohamed, Mohd Nahar Azmi; Jalaludin, Muhammad Yazid

    2016-01-01

    Background: Dietary calcium and vitamin D are essential for bone development. Apart from diet, physical activity may potentially improve and sustain bone health. Objective: To investigate the relationship between the dietary intake of calcium and vitamin D, physical activity, and bone mineral content (BMC) in 13-year-old Malaysian adolescents. Design: Cross-sectional. Setting: Selected public secondary schools from the central and northern regions of Peninsular Malaysia. Participants: The subjects were from the Malaysian Health and Adolescents Longitudinal Research Team Cohort study (MyHeARTs). Methods: The data included seven-day diet histories, anthropometric measurements, and the BMC of calcaneal bone using a portable broadband ultrasound bone densitometer. Nutritionist Pro software was used to calculate the dietary calcium and vitamin D intakes from the diet histories, based on the Nutrient Composition of Malaysian Food Database guidance for the dietary calcium intake and the Singapore Energy and Nutrient Composition of Food Database for vitamin D intake. Results: A total of 289 adolescents (65.7% females) were recruited. The average dietary intakes of calcium and vitamin D were 377 ± 12 mg/day and 2.51 ± 0.12 µg/day, respectively, with the majority of subjects failing to meet the Recommended Nutrient Intake (RNI) of Malaysia for dietary calcium and vitamin D. All the subjects had a normal Z-score for the BMC (−2.00 or higher) with a mean of 0.55 ± 0.01. From the statistical analysis of the factors contributing to BMC, it was found that for those subjects with a higher intake of vitamin D, a higher combination of the intake of vitamin D and calcium resulted in significantly higher BMC quartiles. The regression analysis showed that the BMC might have been influenced by the vitamin D intake. Conclusions: A combination of the intake of vitamin D and calcium is positively associated with the BMC. PMID:27783041

  11. Calcium Imaging of Basal Forebrain Activity during Innate and Learned Behaviors

    PubMed Central

    Harrison, Thomas C.; Pinto, Lucas; Brock, Julien R.; Dan, Yang

    2016-01-01

    The basal forebrain (BF) plays crucial roles in arousal, attention, and memory, and its impairment is associated with a variety of cognitive deficits. The BF consists of cholinergic, GABAergic, and glutamatergic neurons. Electrical or optogenetic stimulation of BF cholinergic neurons enhances cortical processing and behavioral performance, but the natural activity of these cells during behavior is only beginning to be characterized. Even less is known about GABAergic and glutamatergic neurons. Here, we performed microendoscopic calcium imaging of BF neurons as mice engaged in spontaneous behaviors in their home cages (innate) or performed a go/no-go auditory discrimination task (learned). Cholinergic neurons were consistently excited during movement, including running and licking, but GABAergic and glutamatergic neurons exhibited diverse responses. All cell types were activated by overt punishment, either inside or outside of the discrimination task. These findings reveal functional similarities and distinctions between BF cell types during both spontaneous and task-related behaviors. PMID:27242444

  12. Rhinovirus-induced calcium flux triggers NLRP3 and NLRC5 activation in bronchial cells.

    PubMed

    Triantafilou, Kathy; Kar, Satwik; van Kuppeveld, Frank J M; Triantafilou, Martha

    2013-12-01

    Human rhinoviruses have been linked with underlying lung disorders, such as asthma and chronic obstructive pulmonary disease, in children and adults. However, the mechanism of virus-induced airway inflammation is poorly understood. In this study, using virus deletion mutants and silencing for nucleotide-binding oligomerization domain-like receptors (NLRs), we show that the rhinovirus ion channel protein 2B triggers NLRP3 and NLRC5 inflammasome activation and IL-1β secretion in bronchial cells. 2B protein targets the endoplasmic reticulum and Golgi and induces Ca(2+) reduction in these organelles, thereby disturbing the intracellular calcium homeostasis. NLRP3 and NLRC5 act in a cooperative manner during the inflammasome assembly by sensing intracellular Ca(2+) fluxes and trigger IL-1β secretion. These results reveal for the first time that human rhinovirus infection in primary bronchial cells triggers inflammasome activation.

  13. Calcium influx through stretch-activated channels mediates microfilament reorganization in osteoblasts under simulated weightlessness

    NASA Astrophysics Data System (ADS)

    Luo, Mingzhi; Yang, Zhouqi; Li, Jingbao; Xu, Huiyun; Li, Shengsheng; Zhang, Wei; Qian, Airong; Shang, Peng

    2013-06-01

    We have explored the role of Ca2+ signaling in microfilament reorganization of osteoblasts induced by simulated weightlessness using a random positioning machine (RPM). The RPM-induced alterations of cell morphology, microfilament distribution, cell proliferation, cell migration, cytosol free calcium concentration ([Ca2+]i), and protein expression in MG63 osteoblasts were investigated. Simulated weightlessness reduced cell size, disrupted microfilament, inhibited cellular proliferation and migration, and induced an increase in [Ca2+]i in MG63 human osteosarcoma cells. Gadolinium chloride (Gd), an inhibitor for stretch-activated channels, attenuated the increase in [Ca2+]i and microfilament disruption. Further, the expression of calmodulin was significantly increased by simulated weightlessness, and an inhibitor of calmodulin, W-7, aggravated microfilament disruption. Our findings demonstrate that simulated weightlessness induces Ca2+ influx through stretch-activated channels, then results in microfilament disruption.

  14. Cytocompatibility and osteogenic activity of a novel calcium phosphate silicate bioceramic: Silicocarnotite.

    PubMed

    Duan, Wei; Ning, Congqin; Tang, Tingting

    2013-07-01

    In the present study, the effect of a novel bioceramic, silicon-containing calcium phosphate ceramic (silicocarnotite, Ca5 (PO4 )2 SiO4 , CPS) on attachment, proliferation, and osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells (rBMSC) has been investigated in comparison to hydroxyapatite (HA). The CPS showed a similar cell attachment behavior to HA, while the proliferation of rBMSC on CPS was significantly higher than that on HA, which indicated that CPS had a good cytocompatibility. Moreover, the expression of alkaline phosphatase activity and osteogenic-related genes, including Runx-2, osteopontin (OPN), bone sialoprotein (BSP) and osteocalcin (OC), demonstrated that CPS enhanced the osteogenic differentiation of rBMSC and accelerated the differentiation process. The results suggest that CPS ceramic exhibits a good cytocompatibility and osteogenic activity, which might be used as a potential candidate material for bone tissue engineering.

  15. Photocatalytic activity of nanostructured TiO2 films produced by supersonic cluster beam deposition

    NASA Astrophysics Data System (ADS)

    Della Foglia, Flavio; Losco, Tonia; Piseri, Paolo; Milani, Paolo; Selli, Elena

    2009-08-01

    The photocatalytic activity of thin, nanostructured films of titanium dioxide, synthesized by supersonic cluster beam deposition (SCBD) from the gas phase, has been investigated employing the photodegradation of salicylic acid as test reaction. Because of the low deposition energy, the so-deposited highly porous TiO2 films are composed of nanoparticles maintaining their original properties in the film, which can be fully controlled by tuning the deposition and post-deposition treatment conditions. A systematic investigation on the evolution of light absorption properties and photoactivity of the films in relation to their morphology, determined by AFM analysis, and phase composition, determined by Raman spectroscopy, has been performed. The absorption and photocatalytic activity of the nanostructured films in the visible region could be enhanced either through post-deposition annealing treatment in ammonia containing atmosphere or employing mild oxidation conditions, followed by annealing in N2 at 600 °C.

  16. A coarse-grained model to study calcium activation of the cardiac thin filament

    NASA Astrophysics Data System (ADS)

    Zhang, Jing; Schwartz, Steven

    2015-03-01

    Familial hypertrophic cardiomyopathy (FHC) is one of the most common heart disease caused by genetic mutations. Cardiac muscle contraction and relaxation involve regulation of crossbridge binding to the cardiac thin filament, which regulates actomyosin interactions through calcium-dependent alterations in the dynamics of cardiac troponin (cTn) and tropomyosin (Tm). An atomistic model of cTn complex interacting with Tm has been studied by our group. A more realistic model requires the inclusion of the dynamics of actin filament, which is almost 6 times larger than cTn and Tm in terms of atom numbers, and extensive sampling of the model becomes very resource-demanding. By using physics-based protein united-residue force field, we introduce a coarse-grained model to study the calcium activation of the thin filament resulting from cTn's allosteric regulation of Tm dynamics on actin. The time scale is much longer than that of all-atom molecular dynamics simulation because of the reduction of the degrees of freedom. The coarse-grained model is a good template for studying cardiac thin filament mutations that cause FHC, and reduces the cost of computational resources.

  17. Molecular Interactions between Tarantula Toxins and Low-Voltage-Activated Calcium Channels.

    PubMed

    Salari, Autoosa; Vega, Benjamin S; Milescu, Lorin S; Milescu, Mirela

    2016-04-05

    Few gating-modifier toxins have been reported to target low-voltage-activated (LVA) calcium channels, and the structural basis of toxin sensitivity remains incompletely understood. Studies of voltage-gated potassium (Kv) channels have identified the S3b-S4 "paddle motif," which moves at the protein-lipid interface to drive channel opening, as the target for these amphipathic neurotoxins. Voltage-gated calcium (Cav) channels contain four homologous voltage sensor domains, suggesting multiple toxin binding sites. We show here that the S3-S4 segments within Cav3.1 can be transplanted into Kv2.1 to examine their individual contributions to voltage sensing and pharmacology. With these results, we now have a more complete picture of the conserved nature of the paddle motif in all three major voltage-gated ion channel types (Kv, Nav, and Cav). When screened with tarantula toxins, the four paddle sequences display distinct toxin binding properties, demonstrating that gating-modifier toxins can bind to Cav channels in a domain specific fashion. Domain III was the most commonly and strongly targeted, and mutagenesis revealed an acidic residue that is important for toxin binding. We also measured the lipid partitioning strength of all toxins tested and observed a positive correlation with their inhibition of Cav3.1, suggesting a key role for membrane partitioning.

  18. Real-time monitoring of calcium carbonate and cationic peptide deposition on carboxylate-SAM using a microfluidic SAW biosensor

    PubMed Central

    Pohl, Anna

    2014-01-01

    Summary A microfluidic biosensor with surface acoustic wave technology was used in this study to monitor the interaction of calcium carbonate with standard carboxylate self-assembled monolayer sensor chips. Different fluids, with and without biomolecular components, were investigated. The pH-dependent surface interactions of two bio-inspired cationic peptides, AS8 and ES9, which are similar to an extracellular domain of the chitin synthase involved in mollusc shell formation, were also investigated in a biological buffer system. A range of experimental conditions are described that are suitable to study non-covalent molecular interactions in the presence of ionic substances, such as, mineral precursors below the solubility equilibrium. The peptide ES9, equal to the mollusc chitin synthase epitope, is less sensitive to changes in pH than its counterpart AS8 with a penta-lysine core, which lacks the flanking acidic residues. This study demonstrates the extraordinary potential of microfluidic surface acoustic wave biosensors to significantly expand our experimental capabilities for studying the principles underlying biomineralization in vitro. PMID:25383294

  19. Effect of gossypol-acetic acid on calcium transport and ATPase activity in plasma membranes from ram and bull spermatozoa.

    PubMed

    Breitbart, H; Rubinstein, S; Nass-Arden, L

    1984-10-01

    The effects of gossypol acetic acid on the activity of Mg-ATPase and Ca-Mg-ATPase and on calcium uptake by plasma membranes from ram and bull spermatozoa were examined. The three parameters were almost completely inhibited by 10 microM gossypol for both ram and bull sperm. In order to assess the effects of higher gossypol concentrations isolated membrane vesicles were loaded with calcium by operating the ATP-dependent calcium pump after which gossypol was added and calcium uptake followed. At 10 microM gossypol, additional calcium uptake was 85% inhibited while at 40 microM a release of the accumulated calcium was observed. The inhibitory effect of 10 microM gossypol was almost completely reversible by simple dilution of gossypol-treated membranes, whilst at 40 microM the effect was only 50% reversible. The data show a high degree of similarity between bull and ram, suggesting minimal differences between the two species as far as the structure and function of the sperm plasma membrane is concerned.

  20. Differential Effects of G- and F-Actin on the Plasma Membrane Calcium Pump Activity

    PubMed Central

    Vanagas, Laura; de La Fuente, María Candelaria; Dalghi, Marianela; Ferreira-Gomes, Mariela; Rossi, Rolando C.; Strehler, Emanuel E.; Rossi, Juan P. F. C.

    2014-01-01

    We have previously shown that plasma membrane calcium ATPase (PMCA) pump activity is affected by the membrane protein concentration (Vanagas et al., Biochim Biophys Acta 1768:1641–1644, 2007). Results show evidences for the involvement of the actin cytoskeleton. In this study, we explored the relationship between the polymerization state of actin and its effects on purified PMCA activity. Our results show that PMCA associates with the actin cytoskeleton and this interaction causes a modulation of the catalytic activity involving the phosphorylated intermediate of the pump. The state of actin polymerization determines whether it acts as an activator or an inhibitor of the pump: G-actin and/or short oligomers activate the pump, while F-actin inhibits it. The effects of actin on PMCA are the consequence of direct interaction as demonstrated by immunoblotting and cosedimentation experiments. Taken together, these findings suggest that interactions with actin play a dynamic role in the regulation of PMCA-mediated Ca2+ extrusion through the membrane. Our results provide further evidence of the activation–inhibition phenomenon as a property of many cytoskeleton-associated membrane proteins where the cytoskeleton is no longer restricted to a mechanical function but is dynamically involved in modulating the activity of integral proteins with which it interacts. PMID:23152090

  1. Low-Voltage-Activated CaV3.1 Calcium Channels Shape T Helper Cell Cytokine Profiles.

    PubMed

    Wang, Huiyun; Zhang, Xuexin; Xue, Li; Xing, Juan; Jouvin, Marie-Hélène; Putney, James W; Anderson, Matthew P; Trebak, Mohamed; Kinet, Jean-Pierre

    2016-04-19

    Activation of T cells is mediated by the engagement of T cell receptors (TCRs) followed by calcium entry via store-operated calcium channels. Here we have shown an additional route for calcium entry into T cells-through the low-voltage-activated T-type CaV3.1 calcium channel. CaV3.1 mediated a substantial current at resting membrane potentials, and its deficiency had no effect on TCR-initiated calcium entry. Mice deficient for CaV3.1 were resistant to the induction of experimental autoimmune encephalomyelitis and had reduced productions of the granulocyte-macrophage colony-stimulating factor (GM-CSF) by central nervous system (CNS)-infiltrating T helper 1 (Th1) and Th17 cells. CaV3.1 deficiency led to decreased secretion of GM-CSF from in vitro polarized Th1 and Th17 cells. Nuclear translocation of the nuclear factor of activated T cell (NFAT) was also reduced in CaV3.1-deficient T cells. These data provide evidence for T-type channels in immune cells and their potential role in shaping the autoimmune response.

  2. Calcium-dependent modulation by ethanol of mouse synaptosomal pyroglutamyl aminopeptidase activity under basal and K(+)-stimulated conditions.

    PubMed

    Mayas, M D; Ramírez-Expósito, M J; García, M J; Tsuboyama, G; Ramírez, M; Martínez-Martos, J M

    2000-11-03

    We studied the in vitro effects of ethanol (25, 50 and 100 mM) on pyroglutamyl aminopeptidase activity (pGluAP), which has been reported as thyrotrophin-releasing-hormone-degrading activity. pGluAP was measured in presence or absence of calcium, under basal and K(+)-stimulated conditions, in synaptosomes and their incubation supernatant, using pyroglutamyl-beta-naphthylamide as substrate. In basal conditions, in synaptosomes, pGluAP was inhibited by ethanol in a calcium-independent way. In the supernatant, the response differed depending on the concentration of ethanol. Depolarization with K(+) modified pGluAP in synaptosomes and supernatant depending on the presence or not of calcium. In synaptosomes, in absence of calcium, the activity was inhibited at the highest concentrations of ethanol. In contrast, in the supernatant, under depolarizing conditions, ethanol increases pGluAP in absence of calcium. These changes may be in part responsible of the behavioural changes associated to alcohol intake.

  3. Proteolytic maturation of α2δ represents a checkpoint for activation and neuronal trafficking of latent calcium channels

    PubMed Central

    Kadurin, Ivan; Ferron, Laurent; Rothwell, Simon W; Meyer, James O; Douglas, Leon R; Bauer, Claudia S; Lana, Beatrice; Margas, Wojciech; Alexopoulos, Orpheas; Nieto-Rostro, Manuela; Pratt, Wendy S; Dolphin, Annette C

    2016-01-01

    The auxiliary α2δ subunits of voltage-gated calcium channels are extracellular membrane-associated proteins, which are post-translationally cleaved into disulfide-linked polypeptides α2 and δ. We now show, using α2δ constructs containing artificial cleavage sites, that this processing is an essential step permitting voltage-dependent activation of plasma membrane N-type (CaV2.2) calcium channels. Indeed, uncleaved α2δ inhibits native calcium currents in mammalian neurons. By inducing acute cell-surface proteolytic cleavage of α2δ, voltage-dependent activation of channels is promoted, independent from the trafficking role of α2δ. Uncleaved α2δ does not support trafficking of CaV2.2 channel complexes into neuronal processes, and inhibits Ca2+ entry into synaptic boutons, and we can reverse this by controlled intracellular proteolytic cleavage. We propose a model whereby uncleaved α2δ subunits maintain immature calcium channels in an inhibited state. Proteolytic processing of α2δ then permits voltage-dependent activation of the channels, acting as a checkpoint allowing trafficking only of mature calcium channel complexes into neuronal processes. DOI: http://dx.doi.org/10.7554/eLife.21143.001 PMID:27782881

  4. Calcium imaging of motoneuron activity in the en-bloc spinal cord preparation of the neonatal rat.

    PubMed

    Lev-Tov, A; O'Donovan, M J

    1995-09-01

    1. This paper describes the use of calcium imaging to monitor patterns of activity in neonatal rat motoneurons retrogradely labeled with the calcium-sensitive dye, calcium green-dextran. 2. Pressure ejection of calcium green-dextran into ventral roots and into the surgically peeled ventrolateral funiculi (VLF) at the lumbar cord labeled spinal motoneurons and interneurons. The back labeled motoneurons often formed two or three discrete clusters of cells. 3. Fluorescent changes (10-20%) could be detected in labeled motoneurons after a single antidromic stimulus of the segmental ventral root. These changes progressively increased in amplitude during stimulus trains (1-5 s) at frequencies from 5 to 50 Hz, presumably reflecting a frequency-dependent increase in free intracellular calcium. 4. Stimulation of the ipsilateral VLF at the caudal lumbar level (L6), elicited frequency-dependent, synaptically induced motoneuronal discharge. Frequency-dependent fluorescent changes could be detected in calcium green-labeled motoneurons during the VLF-induced synaptic activation. 5. The spatial spread of synaptic activity among calcium green-labeled clusters of motoneurons could be resolved after dorsal root stimulation. Low-intensity stimulation of the roots produced fluorescence changes restricted to the lateral clusters of motoneurons. With increasing stimulation intensity the fluorescence change increased in the lateral cells and could spread into the medial motoneuronal group. After a single supramaximal stimulus a similar pattern was observed with activity beginning laterally and spreading medially. 6. Substantial changes in fluorescence of calcium green-labeled motoneurons were also observed during motoneuron bursting induced by bath application of the glycine receptor antagonist strychnine or the potassium channel blocker 4-aminopyridine (4-AP). 7. Our results show that membrane-impermeant fluorescent calcium indicators can be used as a tool to study the activity of

  5. Eight years of follow-up after laminectomy of calcium pyrophosphate crystal deposition in the cervical yellow ligament of patient with Coffin–Lowry syndrome

    PubMed Central

    Morino, Tadao; Ogata, Tadanori; Horiuchi, Hideki; Yamaoka, Shintaro; Fukuda, Mitsumasa; Miura, Hiromasa

    2016-01-01

    Abstract Background: We report 8 years of follow-up after decompression to treat cervical myelopathy in a patient with Coffin–Lowry syndrome (CLS). CLS is a rare X-linked semidominant syndrome associated with growth and psychomotor retardation, general hypotonia, and skeletal abnormalities. In this patient, the spinal cord was compressed by calcium pyrophosphate crystal deposition in the cervical yellow ligament (YL). To date, only 1 report has described clinical features after surgery for calcified cervical YL in CLS. Methods: A 15-year-old male with tetraplegia secondary to compression of the cervical spinal cord induced by a hypoplastic posterior arch of C1 and calcification of the YL from C2 to C7 was treated surgically with laminectomy from C1 to C7. The patient's history, clinical examination, imaging findings, and treatment are reported. The patient was incapable of speech because of mental retardation, so he could not describe his symptoms. Gait disturbance worsened over the 2 months before admission to our hospital. At admission, the patient could not move his extremities, and tendon reflexes of the upper and lower extremities were significantly increased. Computed tomography of the cervical spine showed YL calcification from C2 to C7. Magnetic resonance imaging showed consecutive compression of the cervical spinal cord. We diagnosed quadriplegia secondary to cervical cord damage and performed emergency surgery. Results: During C1–C7 laminectomy, YL calcification in C2–C7 was observed. The calcification was confirmed as calcium pyrophosphate by crystal analysis. Quadriplegia gradually resolved, and almost disappeared by 2 weeks after the operation. Cervical hyperlordosis was observed in radiographs starting from 1 month after the operation, but it has not progressed and is not associated with any symptoms. Conclusions: The efficacy of decompression continued, and no postoperative complications have occurred during at least 8 years of follow-up. PMID

  6. Inhibition of phospholipase A2 (PLA2) activity by nifedipine and nisoldipine is independent of their calcium-channel-blocking activity

    SciTech Connect

    Chang, J.; Blazek, E.; Carlson, R.P.

    1987-09-01

    The effects of several calcium antagonists on phospholipase A2 (PLA2) activity were examined. Nifedipine and nisoldipine inhibited a cell-free preparation of PLA2 in a dose-dependent manner with maximal inhibition of 71-77% observed at 100 microM. More potent or equipotent dihydropyridine calcium antagonists such as nitrendipine and felodipine did not inhibit PLA2 activity. In addition, nondihydropyridine calcium antagonists such as diltiazem, verapamil, and cinnarazine failed to reduce PLA2 activity markedly. Nifedipine and nisoldipine also reduced PLA2 activity in intact mouse peritoneal macrophages where PLA2 activity was monitored by free (/sup 14/C)arachidonic acid release from (/sup 14/C)arachidonic acid-prelabeled cells. When levels of PGE2 and LTC4 were measured by radioimmunoassay, it was found that the synthesis of these two metabolites was concomitantly inhibited by nifedipine and nisoldipine. In vivo, nifedipine and nisoldipine inhibited tetradecanoylphorbol acetate (TPA) induced ear edema. UV irradiation of nifedipine and nisoldipine (which destroys the slow calcium-channel-blocking activity of these compounds) did not result in a loss of PLA2 inhibitory activity. In fact, in both instances the UV-irradiated forms of nifedipine and nisoldipine were slightly more potent PLA2 inhibitors than the parent compound alone. We therefore conclude that the ability of nifedipine and nisoldipine to inhibit PLA2 was direct and unrelated to their actions on slow calcium channels.

  7. Radiation Dose Deposition in the Active Marrow of Reference Man.

    DTIC Science & Technology

    1977-10-31

    gamma ray-fission neutron exposure , the relative biological effec- tiveness (RBE) per unit marrow dose between neutrons and gamma rays in producing...calculations in terms of marrow dose (rad (marrow)) per unit incident fluence. The third presents in- tegral marrow doses calculated for exposure to...in the marrow than other devices. This is shown by the fact that the neutron dose deposited by a given total exposure from such a de- vice is as much

  8. Shikonin Inhibits Intestinal Calcium-Activated Chloride Channels and Prevents Rotaviral Diarrhea

    PubMed Central

    Jiang, Yu; Yu, Bo; Yang, Hong; Ma, Tonghui

    2016-01-01

    Secretory diarrhea remains a global health burden and causes major mortality in children. There have been some focuses on antidiarrheal therapies that may reduce fluid losses and intestinal motility in diarrheal diseases. In the present study, we identified shikonin as an inhibitor of TMEM16A chloride channel activity using cell-based fluorescent-quenching assay. The IC50 value of shikonin was 6.5 μM. Short-circuit current measurements demonstrated that shikonin inhibited Eact-induced Cl- current in a dose-dependent manner, with IC50 value of 1.5 μM. Short-circuit current measurement showed that shikonin exhibited inhibitory effect against CCh-induced Cl- currents in mouse colonic epithelia but did not affect cytoplasmic Ca2+ concentration as well as the other major enterocyte chloride channel conductance regulator. Characterization study found that shikonin inhibited basolateral K+ channel activity without affecting Na+/K+-ATPase activities. In vivo studies revealed that shikonin significantly delayed intestinal motility in mice and reduced stool water content in a neonatal mice model of rotaviral diarrhea without affecting the viral infection process in vivo. Taken together, the results suggested that shikonin inhibited enterocyte calcium-activated chloride channels, the inhibitory effect was partially through inhbition of basolateral K+ channel activity, and shikonin could be a lead compound in the treatment of rotaviral secretory diarrhea. PMID:27601995

  9. Structural mechanism of ligand activation in human calcium-sensing receptor

    SciTech Connect

    Geng, Yong; Mosyak, Lidia; Kurinov, Igor; Zuo, Hao; Sturchler, Emmanuel; Cheng, Tat Cheung; Subramanyam, Prakash; Brown, Alice P.; Brennan, Sarah C.; Mun, Hee-chang; Bush, Martin; Chen, Yan; Nguyen, Trang X.; Cao, Baohua; Chang, Donald D.; Quick, Matthias; Conigrave, Arthur D.; Colecraft, Henry M.; McDonald, Patricia; Fan, Qing R.

    2016-07-19

    Human calcium-sensing receptor (CaSR) is a G-protein-coupled receptor (GPCR) that maintains extracellular Ca2+homeostasis through the regulation of parathyroid hormone secretion. It functions as a disulfide-tethered homodimer composed of three main domains, the Venus Flytrap module, cysteine-rich domain, and seven-helix transmembrane region. Here, we present the crystal structures of the entire extracellular domain of CaSR in the resting and active conformations. We provide direct evidence that L-amino acids are agonists of the receptor. In the active structure, L-Trp occupies the orthosteric agonist-binding site at the interdomain cleft and is primarily responsible for inducing extracellular domain closure to initiate receptor activation. Our structures reveal multiple binding sites for Ca2+and PO43-ions. Both ions are crucial for structural integrity of the receptor. While Ca2+ions stabilize the active state, PO43-ions reinforce the inactive conformation. The activation mechanism of CaSR involves the formation of a novel dimer interface between subunits.

  10. Structural mechanism of ligand activation in human calcium-sensing receptor

    PubMed Central

    Geng, Yong; Mosyak, Lidia; Kurinov, Igor; Zuo, Hao; Sturchler, Emmanuel; Cheng, Tat Cheung; Subramanyam, Prakash; Brown, Alice P; Brennan, Sarah C; Mun, Hee-chang; Bush, Martin; Chen, Yan; Nguyen, Trang X; Cao, Baohua; Chang, Donald D; Quick, Matthias; Conigrave, Arthur D; Colecraft, Henry M; McDonald, Patricia; Fan, Qing R

    2016-01-01

    Human calcium-sensing receptor (CaSR) is a G-protein-coupled receptor (GPCR) that maintains extracellular Ca2+ homeostasis through the regulation of parathyroid hormone secretion. It functions as a disulfide-tethered homodimer composed of three main domains, the Venus Flytrap module, cysteine-rich domain, and seven-helix transmembrane region. Here, we present the crystal structures of the entire extracellular domain of CaSR in the resting and active conformations. We provide direct evidence that L-amino acids are agonists of the receptor. In the active structure, L-Trp occupies the orthosteric agonist-binding site at the interdomain cleft and is primarily responsible for inducing extracellular domain closure to initiate receptor activation. Our structures reveal multiple binding sites for Ca2+ and PO43- ions. Both ions are crucial for structural integrity of the receptor. While Ca2+ ions stabilize the active state, PO43- ions reinforce the inactive conformation. The activation mechanism of CaSR involves the formation of a novel dimer interface between subunits. DOI: http://dx.doi.org/10.7554/eLife.13662.001 PMID:27434672

  11. Shikonin Inhibits Intestinal Calcium-Activated Chloride Channels and Prevents Rotaviral Diarrhea.

    PubMed

    Jiang, Yu; Yu, Bo; Yang, Hong; Ma, Tonghui

    2016-01-01

    Secretory diarrhea remains a global health burden and causes major mortality in children. There have been some focuses on antidiarrheal therapies that may reduce fluid losses and intestinal motility in diarrheal diseases. In the present study, we identified shikonin as an inhibitor of TMEM16A chloride channel activity using cell-based fluorescent-quenching assay. The IC50 value of shikonin was 6.5 μM. Short-circuit current measurements demonstrated that shikonin inhibited Eact-induced Cl(-) current in a dose-dependent manner, with IC50 value of 1.5 μM. Short-circuit current measurement showed that shikonin exhibited inhibitory effect against CCh-induced Cl(-) currents in mouse colonic epithelia but did not affect cytoplasmic Ca(2+) concentration as well as the other major enterocyte chloride channel conductance regulator. Characterization study found that shikonin inhibited basolateral K(+) channel activity without affecting Na(+)/K(+)-ATPase activities. In vivo studies revealed that shikonin significantly delayed intestinal motility in mice and reduced stool water content in a neonatal mice model of rotaviral diarrhea without affecting the viral infection process in vivo. Taken together, the results suggested that shikonin inhibited enterocyte calcium-activated chloride channels, the inhibitory effect was partially through inhbition of basolateral K(+) channel activity, and shikonin could be a lead compound in the treatment of rotaviral secretory diarrhea.

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

    PubMed Central

    Peng, Yaqin; Liu, Jiane; Miao, Fengqin; Zhang, Jianqiong

    2015-01-01

    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. PMID:26263390

  13. Effects of ethylene glycol tetraacetic acid, A23187 and calmodulin, calcium activated neutral proteinase antagonists on melatonin secretion in perifused chick pineal gland.

    PubMed

    Agapito, M T; Pablos, M; Reiter, R J; Recio, J M; Gutierrez-Baraja, R

    1998-04-10

    We have recently described, using perifused pineal glands, that calcium influx participates in the activation of chick pineal gland. This study shows that the loss of perifused chick pineal gland activity is a complex process which seems to involve the release of calcium from intracellular stores, calmodulin and calcium-activated neutral protease (CANP). Pineal glands were perifused with Krebs medium (controls) or with Krebs medium plus the drugs ethylene glycol tetraacetic acid (EGTA; calcium chelator), A23187 (calcium ionophore), EGTA plus A23187 (extra-intra cellular calcium chelation), trifluoperazine and CGS9343B (calmodulin inhibitors), and E-64 (CANP inhibitor) at the time of the natural peak of melatonin release. When EGTA or A23187 were added to the perifusion medium, no effects were observed. On the other hand, when the calcium chelator EGTA plus A23187 (free extra and intracellular calcium levels were dramatically decreased), trifluoperazine, CGS 9343B or E-64 were added to the perifusion medium melatonin synthesis increased significantly and was sustained for 8 h. We propose a prominent role for calcium output from intracellular stores in regulating melatonin production primarily by acting on Ca-calmodulin and calcium-activated neutral protease.

  14. Structural and functional characterization of a calcium-activated cation channel from Tsukamurella paurometabola

    PubMed Central

    Dhakshnamoorthy, Balasundaresan; Rohaim, Ahmed; Rui, Huan; Blachowicz, Lydia; Roux, Benoît

    2016-01-01

    The selectivity filter is an essential functional element of K+ channels that is highly conserved both in terms of its primary sequence and its three-dimensional structure. Here, we investigate the properties of an ion channel from the Gram-positive bacterium Tsukamurella paurometabola with a selectivity filter formed by an uncommon proline-rich sequence. Electrophysiological recordings show that it is a non-selective cation channel and that its activity depends on Ca2+ concentration. In the crystal structure, the selectivity filter adopts a novel conformation with Ca2+ ions bound within the filter near the pore helix where they are coordinated by backbone oxygen atoms, a recurrent motif found in multiple proteins. The binding of Ca2+ ion in the selectivity filter controls the widening of the pore as shown in crystal structures and in molecular dynamics simulations. The structural, functional and computational data provide a characterization of this calcium-gated cationic channel. PMID:27678077

  15. Structural and functional characterization of a calcium-activated cation channel from Tsukamurella paurometabola

    NASA Astrophysics Data System (ADS)

    Dhakshnamoorthy, Balasundaresan; Rohaim, Ahmed; Rui, Huan; Blachowicz, Lydia; Roux, Benoît

    2016-09-01

    The selectivity filter is an essential functional element of K+ channels that is highly conserved both in terms of its primary sequence and its three-dimensional structure. Here, we investigate the properties of an ion channel from the Gram-positive bacterium Tsukamurella paurometabola with a selectivity filter formed by an uncommon proline-rich sequence. Electrophysiological recordings show that it is a non-selective cation channel and that its activity depends on Ca2+ concentration. In the crystal structure, the selectivity filter adopts a novel conformation with Ca2+ ions bound within the filter near the pore helix where they are coordinated by backbone oxygen atoms, a recurrent motif found in multiple proteins. The binding of Ca2+ ion in the selectivity filter controls the widening of the pore as shown in crystal structures and in molecular dynamics simulations. The structural, functional and computational data provide a characterization of this calcium-gated cationic channel.

  16. Voltage-gated calcium channels function as Ca2+-activated signaling receptors.

    PubMed

    Atlas, Daphne

    2014-02-01

    Voltage-gated calcium channels (VGCCs) are transmembrane cell surface proteins responsible for multifunctional signals. In response to voltage, VGCCs trigger synaptic transmission, drive muscle contraction, and regulate gene expression. Voltage perturbations open VGCCs enabling Ca(2+) binding to the low affinity Ca(2+) binding site of the channel pore. Subsequent to permeation, Ca(2+) targets selective proteins to activate diverse signaling pathways. It is becoming apparent that the Ca(2+)-bound channel triggers secretion in excitable cells and drives contraction in cardiomyocytes prior to Ca(2+) permeation. Here, I highlight recent data implicating receptor-like function of the Ca(2+)-bound channel in converting external Ca(2+) into an intracellular signal. The two sequential mechanistic perspectives of VGCC function are discussed in the context of the prevailing and long-standing current models of depolarization-evoked secretion and cardiac contraction.

  17. Co-immobilization of active antibiotics and cell adhesion peptides on calcium based biomaterials.

    PubMed

    Palchesko, Rachelle N; Buckholtz, Gavin A; Romeo, Jared D; Gawalt, Ellen S

    2014-07-01

    Two bioactive molecules with unrelated functions, vancomycin and a cell adhesion peptide, were immobilized on the surface of a potential bone scaffold material, calcium aluminum oxide. In order to accomplish immobilization and retain bioactivity three sequential surface functionalization strategies were compared: 1.) vancomycin was chemically immobilized before a cell adhesion peptide (KRSR), 2.) vancomycin was chemically immobilized after KRSR and 3.) vancomycin was adsorbed after binding the cell adhesion peptide. Both molecules remained on the surface and active using all three reaction sequences and after autoclave sterilization based on osteoblast attachment, bacterial turbidity and bacterial zone inhibition test results. However, the second strategy was superior at enhancing osteoblast attachment and significantly decreasing bacterial growth when compared to the other sequences.

  18. Glomerular C3c localization indicates ongoing immune deposit formation and complement activation in experimental glomerulonephritis.

    PubMed Central

    Schulze, M.; Pruchno, C. J.; Burns, M.; Baker, P. J.; Johnson, R. J.; Couser, W. G.

    1993-01-01

    In antibody-mediated glomerular disease, deposits of C3 (C3b) are common and are degraded by factor I to C3c and C3d. However, the kinetics of C3b degradation in glomerulonephritis have not been defined. To do this, we studied three models of complement-dependent glomerulonephritis with established C3 deposits (passive Heymann nephritis, cationized immunoglobulin G membranous nephropathy, and concanavalin A-anticoncanavalin A glomerulonephritis). C3b deposition was halted by administration of cobra venom factor, and the disappearance of C3c and C3d from glomeruli was measured with specific antibodies and quantitative fluorescence densitometry. Results showed that C3c deposits were reduced by over 85% within 24 hours in all three models. C3c clearance was unaffected by site or mechanism of deposit formation. C3d deposits persisted despite lack of ongoing complement activation. In passive Heymann nephritis when disease activity was monitored by urinary C5b-9 excretion, C3c was cleared in parallel with return of urine C5b-9 excretion to normal values. We conclude that glomerular deposits of C3c are cleared within 24 hours of cessation of complement activation. Positive staining for C3 utilizing antibody specific for the C3c portion documents recent complement activation usually reflecting new immune deposit formation. Images Figure 1 Figure 2 Figure 3 PMID:7678717

  19. Calcium-deficient apatite: influence of granule size and consolidation mode on release and in vitro activity of vancomycin.

    PubMed

    Obadia, L; Amador, G; Daculsi, G; Bouler, J-M

    2003-03-01

    The use of dynamic compaction and isostatic compression to consolidate calcium phosphate powder loaded with a therapeutic agent avoids a sintering step that could destroy the drug. The present study applied these consolidation methods to vancomycin-loaded calcium-deficient apatite powder, using three granulometric fractions (40-80, 80-200 and 200-500 micrometer). In vitro release profiles were determined via an original system derived from low-pressure liquid chromatography. The biological activity of vancomycin was measured by an in vitro standardized bacteriologic assay, which showed that the drug is completely active after association with calcium phosphate. Regardless of the consolidation method and granulometric fraction used, release profiles were not significantly different and therefore adaptable to injectable suspensions.

  20. Calcium regulation of HCN channels supports persistent activity in a multiscale model of neocortex.

    PubMed

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

    2016-03-01

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

  1. Osteogenic activity of cyclodextrin-encapsulated doxycycline in a calcium phosphate PCL and PLGA composite.

    PubMed

    Trajano, V C C; Costa, K J R; Lanza, C R M; Sinisterra, R D; Cortés, M E

    2016-07-01

    Composites of biodegradable polymers and calcium phosphate are bioactive and flexible, and have been proposed for use in tissue engineering and bone regeneration. When associated with the broad-spectrum antibiotic doxycycline (DOX), they could favor antimicrobial action and enhance the action of osteogenic composites. Composites of polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), and a bioceramic of biphasic calcium phosphate Osteosynt® (BCP) were loaded with DOX encapsulated in β-cyclodextrin (βCD) and were evaluated for effects on osteoblastic cell cultures. The DOX/βCD composite was prepared with a double mixing method. Osteoblast viability was assessed with methyl tetrazolium (MTT) assays after 1day, 7day, and 14days of composite exposure; alkaline phosphatase (AP) activity and collagen production were evaluated after 7days and 14days, and mineral nodule formation after 14days. Composite structures were evaluated by scanning electron microscopy (SEM). Osteoblasts exposed to the composite containing 25μg/mL DOX/βCD had increased cell proliferation (p<0.05) compared to control osteoblast cultures at all experimental time points, reaching a maximum in the second week. AP activity and collagen secretion levels were also elevated in osteoblasts exposed to the DOX/βCD composite (p<0.05 vs. controls) and reached a maximum after 14days. These results were corroborated by Von Kossa test results, which showed strong formation of mineralization nodules during the same time period. SEM of the composite material revealed a surface topography with pore sizes suitable for growing osteoblasts. Together, these results suggest that osteoblasts are viable, proliferative, and osteogenic in the presence of a DOX/βCD-containing BCP ceramic composite.

  2. Evaluation of cytotoxicity, antimicrobial activity and physicochemical properties of a calcium aluminate-based endodontic material

    PubMed Central

    SILVA, Emmanuel João Nogueira Leal; HERRERA, Daniel Rodrigo; ROSA, Tiago Pereira; DUQUE, Thais Mageste; JACINTO, Rogério Castilho; GOMES, Brenda Paula Figueiredo de Almeida; ZAIA, Alexandre Augusto

    2014-01-01

    A calcium aluminate-based endodontic material, EndoBinder, has been developed in order to reduce MTA negative characteristics, preserving its biological properties and clinical applications. Objectives The aim of this study was to evaluate the cytotoxicity, antimicrobial activity, pH, solubility and water sorption of EndoBinder and to compare them with those of white MTA (WMTA). Material and Methods Cytotoxicity was assessed through a multiparametric analysis employing 3T3 cells. Antimicrobial activity against Enterococcus faecalis (ATCC 29212), Staphylococcus aureus. (ATCC 25923) and Candida albicans (ATCC 10556) was determined by the agar diffusion method. pH was measured at periods of 3, 24, 72 and 168 hours. Solubility and water sorption evaluation were performed following ISO requirements. Data were statistically analyzed by ANOVA and Tukey`s test with a significance level of 5%. Results EndoBinder and WMTA were non-cytotoxic in all tested periods and with the different cell viability parameters. There was no statistical differences between both materials (P>.05). All tested materials were inhibitory by direct contact against all microbial strains tested. EndoBinder and WMTA presented alkaline pH in all tested times with higher values of pH for WMTA (P<.05). Both materials showed values complying with the solubility minimum requirements. However, EndoBinder showed lower solubility than WMTA (P<.05). No statistical differences were observed regarding water sorption (P>.05). Conclusion Under these experimental conditions, we concluded that the calcium aluminate-based endodontic material EndoBinder demonstrated suitable biological and physicochemical properties, so it can be suggested as a material of choice in root resorption, perforations and root-end filling. PMID:24626250

  3. Characterization of calcium responses and electrical activity in differentiating mouse neural progenitor cells in vitro.

    PubMed

    de Groot, Martje W G D M; Dingemans, Milou M L; Rus, Katinka H; de Groot, Aart; Westerink, Remco H S

    2014-02-01

    In vitro methods for developmental neurotoxicity (DNT) testing have the potential to reduce animal use and increase insight into cellular and molecular mechanisms underlying chemical-induced alterations in the development of functional neuronal networks. Mouse neural progenitor cells (mNPCs) differentiate into nervous system-specific cell types and have proven valuable to detect DNT using biochemical and morphological techniques. We therefore investigated a number of functional neuronal parameters in primary mNPCs to explore their applicability for neurophysiological in vitro DNT testing. Immunocytochemistry confirmed that mNPCs express neuronal, glial, and progenitor markers at various differentiation durations (1, 7, 14, and 21 days). Because intracellular calcium ([Ca(2+)]i) plays an essential role in neuronal development and function, we measured stimulus-evoked changes in [Ca(2+)]i at these differentiation durations using the Ca(2+)-responsive dye Fura-2. Increases in [Ca(2+)]i (averages ranging from 65 to 226 nM) were evoked by depolarization, ATP, l-glutamic acid, acetylcholine, and dopamine (up to 87%, 57%, 93%, 28%, and 37% responding cells, respectively) and to a lesser extent by serotonin and gamma-aminobutyric acid (both up to 10% responding cells). Notably, the changes in percentage of responsive cells and their response amplitudes over time indicate changes in the expression and functionality of the respective neurotransmitter receptors and related calcium signaling pathways during in vitro differentiation. The development of functional intercellular signaling pathways was confirmed using multielectrode arrays, demonstrating that mNPCs develop electrical activity within 1-2 weeks of differentiation (55% active wells at 14 days of differentiation; mean spike rate of 1.16 spikes/s/electrode). The combined data demonstrate that mNPCs develop functional neuronal characteristics in vitro, making it a promising model to study chemical-induced effects on the

  4. New mechanisms of antiplatelet activity of nifedipine, an L-type calcium channel blocker.

    PubMed

    Chou, Tz-Chong

    2014-01-01

    Platelet hyperactivity often occursd in hypertensive patients and is a key factor in the development of cardiovascular diseases including thrombosis and atherosclerosis. Nifedipine, an L-type calcium channel blocker, is widely used for hypertension and coronary heart disease therapy. In addition, nifedipine is known to exhibit an antiplatelet activity, but the underlying mechanisms involved remain unclear. Several transcription factors such as peroxisome proliferator-activated receptors (PPARs) and nuclear factor kappa B (NF-κB) exist in platelets and have an ability to regulate platelet aggregation through a non-genomic mechanism. The present article focuses on describing the mechanisms of the antiplatelet activity of nifedipine via PPAR activation. It has been demonstrated that nifedipine treatment increases the activity and intracellular amount of PPAR-β/-γ in activated platelets. Moreover, the antiplatelet activity of nifedipine is mediated by PPAR-β/-γ-dependent upon the up-regulation of the PI3K/AKT/NO/cyclic GMP/PKG pathway, and inhibition of protein kinase Cα (PKCα) activity via an interaction between PPAR-β/-γ and PKCα. Furthermore, suppressing NF-κB activation by nifedipine through enhanced association of PPAR-β/-γ with NF-κB has also been observed in collagen-stimulated platelets. Blocking PPAR-β/-γ activity or increasing NF-κB activation greatly reverses the antiplatelet activity and inhibition of intracellular Ca(2+) mobilization, PKCα activity, and surface glycoprotein IIb/IIIa expression caused by nifedipine. Thus, PPAR-β/-γ- dependent suppression of NF-κB activation also contributes to the antiplatelet activity of nifedipine. Consistently, administration of nifedipine markedly reduces fluorescein sodium-induced vessel thrombus formation in mice, which is considerably inhibited when the PPAR-β/-γ antagonists are administrated simultaneously. Collectively, these results provide important information regarding the mechanism by

  5. Selective regulation of spontaneous activity of neurons of the deep cerebellar nuclei by N-type calcium channels in juvenile rats.

    PubMed

    Alviña, Karina; Khodakhah, Kamran

    2008-05-15

    The cerebellum coordinates movement and maintains body posture. The main output of the cerebellum is formed by three deep nuclei, which receive direct inhibitory inputs from cerebellar Purkinje cells, and excitatory collaterals from mossy and climbing fibres. Neurons of deep cerebellar nuclei (DCN) are spontaneously active, and disrupting their activity results in severe cerebellar ataxia. It is suggested that voltage-gated calcium channels make a significant contribution to the spontaneous activity of DCN neurons, although the exact identity of these channels is not known. We sought to delineate the functional role and identity of calcium channels that contribute to pacemaking in DCN neurons of juvenile rats. We found that in the majority of cells blockade of calcium currents results in avid high-frequency bursting, consistent with the notion that the net calcium-dependent current in DCN neurons is outward. We showed that the bursting seen in these neurons after block of calcium channels is the consequence of reduced activation of small-conductance calcium-activated (SK) potassium channels. With the use of selective pharmacological blockers we showed that L-, P/Q-, R- and T-type calcium channels do not contribute to the spontaneous activity of DCN neurons. In contrast, blockade of high-threshold N-type calcium channels increased the firing rate and caused the cells to burst. Our results thus suggest a selective coupling of N-type voltage-gated calcium channels with calcium-activated potassium channels in DCN neurons. In addition, we demonstrate the presence of a cadmium-sensitive calcium conductance coupled with SK channels, that is pharmacologically distinct from L-, N-, P/Q-, R- and T-type calcium channels.

  6. Calcium/calmodulin kinase II activity of hippocampus in kainate-induced epilepsy.

    PubMed Central

    Lee, M. C.; Ban, S. S.; Woo, Y. J.; Kim, S. U.

    2001-01-01

    This study investigated calcium/calmodulin kinase II (CaMKII) activity related to long-standing neuronal injury of the hippocampus in kainate (KA)-induced experimental temporal lobe epilepsy. Epileptic seizure was induced by injection of KA (1 microg/microL) dissolved in phosphate buffer (0.1 M, pH 7.4) into the left amygdala. Clinical seizures, histopathologic changes and CaMKII activity of the hippocampus were evaluated. Characteristic early limbic and late seizures were developed. Hippocampal CaMKII activity increased significantly 4 and 8 weeks after intra-amygdaloid injection of KA, when late seizures developed. The histopathologic changes of the hippocampus included swelling of neuronal cytoplasm with nuclear pyknosis and loss of neurons in CA3 during this period. The increased activity of CaMKII may correlate with appearance of distant damage in the hippocampus. The above results indicate that intra-amygdaloid injection of KA produces excitatory signals for ipsilateral CA3 neurons in the hippocampus and that subsequently increased levels of CaMKII in postsynaptic neurons induce neuronal injury via phosphorylation of N-methyl-D-aspartate type glutamate receptor. PMID:11641537

  7. Calcium influx-mediated signaling is required for complete mouse egg activation.

    PubMed

    Miao, Yi-Liang; Stein, Paula; Jefferson, Wendy N; Padilla-Banks, Elizabeth; Williams, Carmen J

    2012-03-13

    Mammalian fertilization is accompanied by oscillations in egg cytoplasmic calcium (Ca(2+)) concentrations that are critical for completion of egg activation. These oscillations are initiated by Ca(2+) release from inositol 1,4,5-trisphosphate (IP(3))-sensitive intracellular stores. We tested the hypothesis that Ca(2+) influx across the plasma membrane was a requisite component of egg activation signaling, and not simply a Ca(2+) source for store repletion. Using intracytoplasmic sperm injection (ICSI) and standard in vitro fertilization (IVF), we found that Ca(2+) influx was not required to initiate resumption of meiosis II. However, even if multiple oscillations in intracellular Ca(2+) occurred, in the absence of Ca(2+) influx, the fertilized eggs failed to emit the second polar body, resulting in formation of three pronuclei. Additional experiments using the Ca(2+) chelator, BAPTA/AM, demonstrated that Ca(2+) influx is sufficient to support polar body emission and pronucleus formation after only a single sperm-induced Ca(2+) transient, whereas BAPTA/AM-treated ICSI or fertilized eggs cultured in Ca(2+)-free medium remained arrested in metaphase II. Inhibition of store-operated Ca(2+) entry had no effect on ICSI-induced egg activation, so Ca(2+) influx through alternative channels must participate in egg activation signaling. Ca(2+) influx appears to be upstream of CaMKIIγ activity because eggs can be parthenogenetically activated with a constitutively active form of CaMKIIγ in the absence of extracellular Ca(2+). These results suggest that Ca(2+) influx at fertilization not only maintains Ca(2+) oscillations by replenishing Ca(2+) stores, but also activates critical signaling pathways upstream of CaMKIIγ that are required for second polar body emission.

  8. Chronic imaging of movement-related Purkinje cell calcium activity in awake behaving mice

    PubMed Central

    Gaffield, Michael A.; Amat, Samantha B.; Bito, Haruhiko

    2015-01-01

    Purkinje cells (PCs) are a major site of information integration and plasticity in the cerebellum, a brain region involved in motor task refinement. Thus PCs provide an ideal location for studying the mechanisms necessary for cerebellum-dependent motor learning. Increasingly, sophisticated behavior tasks, used in combination with genetic reporters and effectors of activity, have opened up the possibility of studying cerebellar circuits during voluntary movement at an unprecedented level of quantitation. However, current methods used to monitor PC activity do not take full advantage of these advances. For example, single-unit or multiunit electrode recordings, which provide excellent temporal information regarding electrical activity, only monitor a small population of cells and can be quite invasive. Bolus loading of cell-permeant calcium (Ca2+) indicators is short-lived, requiring same-day imaging immediately after surgery and/or indicator injection. Genetically encoded Ca2+ indicators (GECIs) overcome many of these limits and have garnered considerable use in many neuron types but only limited use in PCs. Here we employed these indicators to monitor Ca2+ activity in PCs over several weeks. We could repeatedly image from the same cerebellar regions across multiple days and observed stable activity. We used chronic imaging to monitor PC activity in crus II, an area previously linked to licking behavior, and identified a region of increased activity at the onset of licking. We then monitored this same region after training tasks to initiate voluntary licking behavior in response to different sensory stimuli. In all cases, PC Ca2+ activity increased at the onset of rhythmic licking. PMID:26561609

  9. The prevalence of chondrocalcinosis (CC) of the acromioclavicular (AC) joint on chest radiographs and correlation with calcium pyrophosphate dihydrate (CPPD) crystal deposition disease.

    PubMed

    Parperis, Konstantinos; Carrera, Guillermo; Baynes, Keith; Mautz, Alan; Dubois, Melissa; Cerniglia, Ross; Ryan, Lawrence M

    2013-09-01

    Digital imaging combined with picture archiving and communication system (PACS) access allows detailed image retrieval and magnification. Calcium pyrophosphate dihydrate (CPPD) crystals preferentially deposit in fibrocartilages, the cartilage of the acromioclavicular (AC) joint being one such structure. We sought to determine if examination of the AC joints on magnified PACS imaging of chest films would be useful in identifying chondrocalcinosis (CC). Retrospective radiographic readings and chart reviews involving 1,920 patients aged 50 or more who had routine outpatient chest radiographs over a 4-month period were performed. Knee radiographs were available for comparison in 489 patients. Medical records were reviewed to abstract demographics, chest film reports, and diagnoses. AC joint CC was identified in 1.1 % (21/1,920) of consecutive chest films. Patients with AC joint CC were 75 years of age versus 65.4 in those without CC (p < 0.0002). Four hundred eighty-nine patients had knee films. Six of these patients had AC joint CC, and of these, five also had knee CC (83 %). Of the 483 without AC joint CC, 62 (12 %) had knee CC (p = 0.002). Patients with AC joint CC were more likely to have a recorded history of CPPD crystal deposition disease than those without AC joint CC (14 versus 1 %, p = 0.0017). The prevalence of AC joint CC increases with age and is associated with knee CC. A finding of AC joint CC should heighten suspicion of pseudogout or secondary osteoarthritis in appropriate clinical settings and, in a young patient, should alert the clinician to the possibility of an associated metabolic condition.

  10. Electrospray deposition in vacuum as method to create functionally active protein immobilization on polymeric substrates.

    PubMed

    Fornari, Enzo; Roberts, Clive J; Temperton, Robert H; O'Shea, James N

    2015-09-01

    We demonstrate in this work the deposition of a large biological molecule (fibronectin) on polymeric substrates in a high vacuum environment using an electrospray deposition system. Fibronectin was deposited and its distribution and structure investigated and retention of function (ability to promote cell adhesion) on return to liquid environment is shown. AFM was used to monitor changes in the morphology of the surface before and after fibronectin deposition, whilst the biological activity of the deposited protein is assessed through a quantitative analysis of the biomolecular adhesion and migration of fibroblast cells to the modified surfaces. For the first time we have demonstrated that using high vacuum electrospray deposition it is possible to deposit large protein molecules on polymeric surfaces whilst maintaining the protein activity. The deposition of biological molecules such as proteins with the retention of their activity onto clean well-controlled surfaces under vacuum condition, offers the possibility for future studies utilizing high resolution vacuum based techniques at the atomic and molecular scale providing a greater understanding of protein-surface interface behaviour of relevance to a wide range of applications such as in sensors, diagnostics and tissue engineering.

  11. EFFECTS OF SODIUM AND CALCIUM IN LIGNITE ON THE PERFORMANCE OF ACTIVATED CARBON PRODUCTS

    SciTech Connect

    Edwin S. Olson; Kurt E. Eylands; Daniel J. Stepan

    2001-12-01

    New federal drinking water regulations have been promulgated to restrict the levels of disinfection by-products (DBPs) in finished public water supplies. DBPs are suspected carcinogens and are formed when organic material is partially oxidized by disinfectants commonly used in the water treatment industry. Additional federal mandates are expected in the near future that will also affect public water suppliers with respect to DBPs. These new federal drinking water regulations may require public water suppliers to adjust treatment practices or incorporate additional treatment operations into their existing treatment trains. Many options have been identified, including membrane processes, granular activated carbon, powered activated carbon (PAC), enhanced coagulation and/or softening, and alternative disinfectants (e.g., chlorine dioxide, ozone, and chloramines). Of the processes being considered, PAC appears to offer an attractive benefit-to-cost advantage for many water treatment plants, particularly small systems (those serving fewer than 10,000 customers). PAC has traditionally been used by the water treatment industry for the removal of compounds contributing to taste and odor problems. PAC also has the potential to remove naturally occurring organic matter (NOM) from raw waters prior to disinfection, thus controlling the formation of regulated DBPs. Many small water systems are currently using PAC for taste and odor control and have the potential to use PAC for controlling DBPs. Activated carbons can be produced from a variety of raw materials, including wood, peat, coconut husks, and numerous types of coal. The Energy & Environmental Research Center (EERC) has been working on the development of a PAC product to remove NOM from surface water supplies to prevent the formation of carcinogenic DBPs during chlorination. During that study, the sodium and calcium content of the lignites showed a significant effect on the sorption capacity of the activated carbon

  12. Potassium permeability activated by intracellular calcium ion concentration in the pancreatic beta-cell.

    PubMed Central

    Atwater, I; Dawson, C M; Ribalet, B; Rojas, E

    1979-01-01

    1. Membrane potentials and input resistance were measured in beta-cells from mouse pancreatic islets of Langerhans in a study designed to assess the role of a K permeability specifically blocked by quinine or quinidine and activated by intracellular calcium ion concentration ([Ca2+])i-activated PK). 2. Addition of 100 microM-quinine to the perifusion medium resulted in a 10--30 mV depolarization of the membrane and an increase in the input resistance of ca. 4.10(7) omega. 3. In the absence of glucose, 100 microM-quinine induced electrical activity. 4. In the presence of glucose, 100 microM-quinine abolished the burst pattern of electrical activity and very much reduced the graded response of spike frequency normally seen with different concentrations of glucose. 5. Addition of mitochondrial inhibitors, KCN, NaN3, DNP, CCCP, FCCP, to the perifusion medium containing glucose rapidly hyperpolarized the beta-cell membrane, inducing a concomitant decrease in input resistance. 6. In the presence of glucose, these mitochondrial inhibitors reversibly blocked electrical activity; upon removal of the inhibitor, recovery of electrical activity followed a biphasic pattern. 7. The effects of mitochondrial inhibitors were partially reversed by 100 microM-quinine. 8. It is proposed that the membrane potential of the beta-cell in the absence of glucose is predominantly controlled by the [Ca2+]i-activated PK. It is further suggested that this permeability to K controls the level for glucose stimulation and leads to the generation of the burst pattern. PMID:381636

  13. Inhibition of Glycolate Oxidase With Dicer-substrate siRNA Reduces Calcium Oxalate Deposition in a Mouse Model of Primary Hyperoxaluria Type 1

    PubMed Central

    Dutta, Chaitali; Avitahl-Curtis, Nicole; Pursell, Natalie; Larsson Cohen, Marita; Holmes, Benjamin; Diwanji, Rohan; Zhou, Wei; Apponi, Luciano; Koser, Martin; Ying, Bo; Chen, Dongyu; Shui, Xue; Saxena, Utsav; Cyr, Wendy A; Shah, Anee; Nazef, Naim; Wang, Weimin; Abrams, Marc; Dudek, Henryk; Salido, Eduardo; Brown, Bob D; Lai, Chengjung

    2016-01-01

    Primary hyperoxaluria type 1 (PH1) is an autosomal recessive, metabolic disorder caused by mutations of alanine-glyoxylate aminotransferase (AGT), a key hepatic enzyme in the detoxification of glyoxylate arising from multiple normal metabolic pathways to glycine. Accumulation of glyoxylate, a precursor of oxalate, leads to the overproduction of oxalate in the liver, which accumulates to high levels in kidneys and urine. Crystalization of calcium oxalate (CaOx) in the kidney ultimately results in renal failure. Currently, the only treatment effective in reduction of oxalate production in patients who do not respond to high-dose vitamin B6 therapy is a combined liver/kidney transplant. We explored an alternative approach to prevent glyoxylate production using Dicer-substrate small interfering RNAs (DsiRNAs) targeting hydroxyacid oxidase 1 (HAO1) mRNA which encodes glycolate oxidase (GO), to reduce the hepatic conversion of glycolate to glyoxylate. This approach efficiently reduces GO mRNA and protein in the livers of mice and nonhuman primates. Reduction of hepatic GO leads to normalization of urine oxalate levels and reduces CaOx deposition in a preclinical mouse model of PH1. Our results support the use of DsiRNA to reduce liver GO levels as a potential therapeutic approach to treat PH1. PMID:26758691

  14. EFFECT OF ACTIVE ACCUMULATION OF CALCIUM AND PHOSPHATE IONS ON THE STRUCTURE OF RAT LIVER MITOCHONDRIA.

    PubMed

    GREENAWALT, J W; ROSSI, C S; LEHNINGER, A L

    1964-10-01

    Rat liver mitochondria allowed to accumulate maximal amounts of Ca(++) and HPO(4) (=) ions from the suspending medium in vitro during respiration have a considerably higher specific gravity than normal mitochondria and may be easily separated from the latter by isopycnic centrifugation in density gradients of sucrose or cesium chloride. When the mitochondria are allowed to accumulate less than maximal amounts of Ca(++) and HPO(4) (=) from the medium, they have intermediate specific gravities which are roughly proportional to their content of calcium phosphate. Maximally "loaded" mitochondria are relatively homogeneous with respect to specific gravity. Correlated biochemical and electron microscopic studies show that Ca(++)-loaded mitochondria contain numerous dense granules, of which some 85 per cent are over 500 A in diameter. These granules are electron-opaque not only following fixation and staining with heavy metal reagents, but also following fixation with formaldehyde, demonstrating that the characteristic granules in Ca(++)-loaded mitochondria have intrinsic electron-opacity. The dense granules are almost always located within the inner compartment of the mitochondria and not in the space between the inner and outer membranes. They are frequently located at or near the cristae and they often show electron-transparent "cores." Such granules appear to be made up of clusters of smaller dense particles, but preliminary x-ray diffraction analysis and electron diffraction studies have revealed no evidence of crystallinity in the deposits. The electron-opaque granules decrease in number when the Ca(++)-loaded mitochondria are incubated with 2,4-dinitrophenol; simultaneously there is discharge of Ca(++) and phosphate from the mitochondria into the medium.

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

    PubMed

    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.

  16. Sonic hedgehog stimulates the proliferation of rat gastric mucosal cells through ERK activation by elevating intracellular calcium concentration

    SciTech Connect

    Osawa, Hiroyuki; Ohnishi, Hirohide . E-mail: hohnishi@jichi.ac.jp; Takano, Koji; Noguti, Takasi; Mashima, Hirosato; Hoshino, Hiroko; Kita, Hiroto; Sato, Kiichi; Matsui, Hirofumi; Sugano, Kentaro

    2006-06-02

    Sonic Hedgehog (Shh), a member of hedgehog peptides family, is expressed in gastric gland epithelium. To elucidate Shh function to gastric mucosal cells, we examined the effect of Shh on the proliferation of a rat normal gastric mucosal cell line, RGM-1. RGM-1 cells express essential components of Shh receptor system, patched-1, and smoothened. Shh enhanced DNA synthesis in RGM-1 cells and elevated intracellular calcium concentration ([Ca{sup 2+}]{sub i}). In addition, Shh as well as calcium ionophore A32187 rapidly activated ERK. However, Shh failed to activate ERK under calcium-free culture condition. Pretreatment of cells with PD98059 attenuated the DNA synthesis promoted by Shh. Moreover, when cells were pretreated with cyclopamine, Shh could not elevate [Ca{sup 2+}]{sub i}, activate ERK or promote DNA synthesis. On the other hand, although Shh induced Gli-1 nuclear accumulation in RGM-1 cells, Shh activated ERK even in cells pretreated with actinomycin D. These results indicate that Shh promotes the proliferation of RGM-1 cells through an intracellular calcium- and ERK-dependent but transcription-independent pathway via Patched/Smoothened receptor system.

  17. Sources of activator calcium for potassium- and serotonin-induced constriction of isolated bovine cerebral arteries

    SciTech Connect

    Not Available

    1986-03-01

    Previous in vitro studies with the calcium channel blockers (CCB) indirectly suggest that K/sup +/ and serotonin (5HT) constrict bovine middle cerebral arteries (BMCA) by promoting the influx of extracellular calcium (Ca) through CCB-sensitive channels. In this study, the authors directly determined the sources of activator Ca for K/sup +/- and 5HT-induced constriction of BMCA, using radiolabelled /sup 4/)2%Ca and /sup 3/H-sorbitol. EGTA-resistant Ca uptake, an estimate of Ca influx into vascular smooth muscle, was determined by exposure to Ca-deficient 2 mM EGTA solutions at 1/sup 0/C. The total Ca content of BMCA was 4.4 nmole/mg (wet wt.) after equilibration at 37/sup 0/C. The total exchangeable Ca content was 1.64 nmole/mg after 1 hr of /sup 45/Ca loading; the Ca content of the extracellular water was 0.30 nmole/mg, as estimated from the /sup 3/H-sorbitol space (0.25 ul/mg). The EGTA-resistant Ca uptake at 1 hr was 134 pmole/mg. K/sup +/ and 5HT significantly increased EGTA-resistant Ca uptake during 5 min of /sup 45/Ca loading; for K/sup +/, Ca uptake increased from 71 to 202 pmole/mg, and for 5HT, from 65 to 102 pmole/mg. Verapamil (10/sup -5/ M) or nifedipine (3.3 x 10/sup -7/ M) significantly blocked the increase in EGTA-resistant Ca uptake induced by K/sup +/ or 5HT. These results provide direct evidence that K/sup +/ or 5HT may constrict BMCA by promoting the influx of extracellular Ca through CCB-sensitive channels.

  18. Calcium ion gradients modulate the zinc affinity and antibacterial activity of human calprotectin.

    PubMed

    Brophy, Megan Brunjes; Hayden, Joshua A; Nolan, Elizabeth M

    2012-10-31

    Calprotectin (CP) is an antimicrobial protein produced and released by neutrophils that inhibits the growth of pathogenic microorganisms by sequestering essential metal nutrients in the extracellular space. In this work, spectroscopic and thermodynamic metal-binding studies are presented to delineate the zinc-binding properties of CP. Unique optical absorption and EPR spectroscopic signatures for the interfacial His(3)Asp and His(4) sites of human calprotectin are identified by using Co(II) as a spectroscopic probe. Zinc competition titrations employing chromophoric Zn(II) indicators provide a 2:1 Zn(II):CP stoichiometry, confirm that the His(3)Asp and His(4) sites of CP coordinate Zn(II), and reveal that the Zn(II) affinity of both sites is calcium-dependent. The calcium-insensitive Zn(II) competitor ZP4 affords dissociation constants of K(d1) = 133 ± 58 pM and K(d2) = 185 ± 219 nM for CP in the absence of Ca(II). These values decrease to K(d1) ≤ 10 pM and K(d2) ≤ 240 pM in the presence of excess Ca(II). The K(d1) and K(d2) values are assigned to the His(3)Asp and His(4) sites, respectively. In vitro antibacterial activity assays indicate that the metal-binding sites and Ca(II)-replete conditions are required for CP to inhibit the growth of both Gram-negative and -positive bacteria. Taken together, these data provide a working model whereby calprotectin responds to physiological Ca(II) gradients to become a potent Zn(II) chelator in the extracellular space.

  19. Physicochemical properties and osteogenic activity of radiopaque calcium silicate-gelatin cements.

    PubMed

    Wang, Chien-Wen; Chiang, Ting-Yi; Chang, Hsien-Chang; Ding, Shinn-Jyh

    2014-09-01

    The purpose of this study is to evaluate the physicochemical properties and in vitro osteogenic activity of radiopaque calcium silicate-gelatin cements. The radiopacity, setting time, working time, flow, diametral tensile strength, pH value, washout resistance and morphology of the cements with gelatin (0, 5 and 10% by weight) were measured, which compared to a popular endodontic material, ProRoot white-colored mineral trioxide aggregate (WMTA). The cell morphology, cell attachment and proliferation, alkaline phosphatase and osteocalcin levels on the cements were measured by culturing the specimens with dental pulp cells. The results indicated that the presence of gelatin significantly (P < 0.05) reduced radiopacity and diametral tensile strength and prolonged setting time. Nevertheless, the 5 wt% gelatin cement had a radiopacity (5.1 mm of Al thickness) higher than ISO 6876:2001 standards (3 mm of Al thickness). The setting time (33 min), working time (9 min) and flow value (17.4 mm) of the 5 wt% gelatin cement were significantly (P < 0.05) better than those of WMTA (corresponding 165, 6 min and 14.2 mm). The fresh WMTA completely degraded after soaking in a physiological solution for 1 h, while the gelatin cements resisted washout, showing no noticeable breakdown even after 1 day of soaking. The gelatin cement enhanced the higher expression of cell attachment, proliferation and differentiation as compared to WMTA. It was concluded that the 5 wt% gelatin-calcium silicate hybrid cement appears to be promising as a radiopaque biomaterial for medical applications such as endodontics and vertebroplasty.

  20. Serotonin Regulates Calcium Homeostasis in Lactation by Epigenetic Activation of Hedgehog Signaling

    PubMed Central

    Laporta, Jimena; Keil, Kimberly P.; Weaver, Samantha R.; Cronick, Callyssa M.; Prichard, Austin P.; Crenshaw, Thomas D.; Heyne, Galen W.; Vezina, Chad M.; Lipinski, Robert J.

    2014-01-01

    Calcium homeostasis during lactation is critical for maternal and neonatal health. We previously showed that nonneuronal/peripheral serotonin [5-hydroxytryptamine (5-HT)] causes the lactating mammary gland to synthesize and secrete PTHrP in an acute fashion. Here, using a mouse model, we found that genetic inactivation of tryptophan hydroxylase 1 (Tph1), which catalyzes the rate-limiting step in peripheral 5-HT synthesis, reduced circulating and mammary PTHrP expression, osteoclast activity, and maternal circulating calcium concentrations during the transition from pregnancy to lactation. Tph1 inactivation also reduced sonic hedgehog signaling in the mammary gland during lactation. Each of these deficiencies was rescued by daily injections of 5-hydroxy-L-tryptophan (an immediate precursor of 5-HT) to Tph1-deficient dams. We used immortalized mouse embryonic fibroblasts to demonstrate that 5-HT induces PTHrP through a sonic hedgehog-dependent signal transduction mechanism. We also found that 5-HT altered DNA methylation of the Shh gene locus, leading to transcriptional initiation at an alternate start site and formation of a variant transcript in mouse embryonic fibroblasts in vitro and in mammary tissue in vivo. These results support a new paradigm of 5-HT-mediated Shh regulation involving DNA methylation remodeling and promoter switching. In addition to having immediate implications for lactation biology, identification and characterization of a novel functional regulatory relationship between nonneuronal 5-HT, hedgehog signaling, and PTHrP offers new avenues for the study of these important factors in development and disease. PMID:25192038

  1. Calcium signals activated by ghrelin and D-Lys(3)-GHRP-6 ghrelin antagonist in developing dorsal root ganglion glial cells.

    PubMed

    Erriquez, Jessica; Bernascone, Silvia; Ciarletta, Monica; Filigheddu, Nicoletta; Graziani, Andrea; Distasi, Carla

    2009-09-01

    Ghrelin is a hormone regulating energy homeostasis via interaction with its receptor, GHSR-1a. Ghrelin activities in dorsal root ganglia (DRG) cells are unknown. Herein we show that ghrelin induces a change of cytosolic calcium concentration in both glia and neurons of embryonic chick DRG. Both RT-PCR and binding studies performed with fluorescent ghrelin in the presence of either unlabeled ghrelin or GHSR-1a antagonist D-Lys(3)-GHRP-6, indicate that DRG cells express GHSR-1a. In glial cells the response is characterized by a rapid transient rise in [Ca(2+)](i) followed by a long lasting rise. The calcium elevation is dependent on calcium release from thapsigargin-sensitive intracellular stores and on activation of two distinct Ca(2+) entry pathways, a receptor activated calcium entry and a store operated calcium entry. Surprisingly, D-Lys(3)-GHRP-6 exerts several activities in the absence of exogenous ghrelin: (i) it activates calcium release from thapsigargin-sensitive intracellular stores and calcium entry via voltage-operated channels in non-neuronal cells; (ii) it inhibits calcium oscillations in non-neuronal cells exhibiting spontaneous Ca(2+) activity and iii) it promotes apoptosis of DRG cells, both neurons and glia. In summary, we provide the first evidence for ghrelin activity in DRG, and we also demonstrate that the widely used D-Lys(3)-GHRP-6 ghrelin antagonist features ghrelin independent activities.

  2. Redistribution of cathepsin B activity from the endosomal-lysosomal pathway in chick intestine within 3 min of calcium absorption.

    PubMed

    Nemere, I; Norman, A W

    1991-06-01

    Earlier work has suggested that calcium-containing lysosomes are involved in 1,25-dihydroxyvitamin D3 (1,25(OH)2D3)-stimulated intestinal absorption of the divalent cation. In the present report immunofluorescent labelling studies on fixed frozen sections of chick intestine were undertaken to determine whether lysosomes could respond to calcium transport conditions in less than 5 min. Tissue prepared from vitamin D-deficient chicks dosed with vehicle or 1.3 nmol of 1,25(OH)2D3 15 h prior to use was immunofluorescently labelled for cathepsin B, a lysosomal protease. In the absence of calcium absorption, punctate staining was found in the region below the terminal web, and more diffusely in the cytoplasm. The intensity of staining was noticeably greater in sections from 1,25(OH)2D3-treated than control chicks. In sections prepared after 3 min of calcium absorption, cathepsin B staining was localized near the basal and lateral membranes of the epithelial cells. After 30 min of transport, the protease was found in the villus core regardless of vitamin D status; however, immunoreactivity within the epithelial cells of 1,25(OH)2D3-treated chick intestine had returned to pretransport intensity, whereas that of controls had not. To further investigate the specificity of the cathepsin B antibody, the intracellular compartmentalization of the protease was determined by biochemical methods. Using dosing procedures and calcium transport times equivalent to those for the immunofluorescent studies mucosae were collected by scraping, homogenized, and subcellular fractions prepared by a combination of differential and Percoll gradient centrifugation. In the absence of calcium transport, cathepsin B-specific activity was enhanced in whole homogenates, endocytic vesicles, and a lysosomal fraction prepared from intestinal epithelium of 1,25(OH)2D3-treated chicks, relative to vitamin D-deficient controls. After 3 min of calcium absorption, a profound (approximately 4-fold) decrease in

  3. Degradation of trichloroethylene in aqueous solution by calcium peroxide activated with ferrous ion.

    PubMed

    Zhang, Xiang; Gu, Xiaogang; Lu, Shuguang; Miao, Zhouwei; Xu, Minhui; Fu, Xiaori; Qiu, Zhaofu; Sui, Qian

    2015-03-02

    The application of calcium peroxide (CaO2) activated with ferrous ion to stimulate the degradation of trichloroethylene (TCE) was investigated. The experimental results showed that TCE could be completely degraded in 5 min at a CaO2/Fe(II)/TCE molar ratio of 4/8/1. Probe compound tests demonstrated the presence of reactive oxygen species HO· and O2(-·) in CaO2/Fe(II) system, while scavenging tests indicated that HO· was the dominant active species responsible for TCE removal, and O2(-·) could promote TCE degradation in CaO2/Fe(II) system. In addition, the influences of initial solution pH and solution matrix were evaluated. It suggested that the elevation of initial solution pH suppressed TCE degradation. Cl(-) had significant scavenging effect on TCE removal, whereas HCO3(-) of high concentration showed favorable function. The influences of NO3(-) and SO4(2-) could be negligible, while natural organic matter (NOM) had a negative effect on TCE removal at a relatively high concentration. The results demonstrated that the technique of CaO2 activated with ferrous ion is a highly promising technique in in situ chemical oxidation (ISCO) remediation in TCE contaminated sites.

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

    PubMed

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

    2015-04-14

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

  5. Complete Atrial-Specific Knockout of Sodium-Calcium Exchange Eliminates Sinoatrial Node Pacemaker Activity

    PubMed Central

    Groenke, Sabine; Larson, Eric D.; Alber, Sarah; Zhang, Rui; Lamp, Scott T.; Ren, Xiaoyan; Nakano, Haruko; Jordan, Maria C.; Karagueuzian, Hrayr S.; Roos, Kenneth P.; Nakano, Atsushi; Proenza, Catherine; Philipson, Kenneth D.; Goldhaber, Joshua I.

    2013-01-01

    The origin of sinoatrial node (SAN) pacemaker activity in the heart is controversial. The leading candidates are diastolic depolarization by “funny” current (If) through HCN4 channels (the “Membrane Clock“ hypothesis), depolarization by cardiac Na-Ca exchange (NCX1) in response to intracellular Ca cycling (the "Calcium Clock" hypothesis), and a combination of the two (“Coupled Clock”). To address this controversy, we used Cre/loxP technology to generate atrial-specific NCX1 KO mice. NCX1 protein was undetectable in KO atrial tissue, including the SAN. Surface ECG and intracardiac electrograms showed no atrial depolarization and a slow junctional escape rhythm in KO that responded appropriately to β-adrenergic and muscarinic stimulation. Although KO atria were quiescent they could be stimulated by external pacing suggesting that electrical coupling between cells remained intact. Despite normal electrophysiological properties of If in isolated patch clamped KO SAN cells, pacemaker activity was absent. Recurring Ca sparks were present in all KO SAN cells, suggesting that Ca cycling persists but is uncoupled from the sarcolemma. We conclude that NCX1 is required for normal pacemaker activity in murine SAN. PMID:24278453

  6. Dynamic calcium requirements for activation of rabbit papillary muscle calculated from tension-independent heat.

    PubMed

    Blanchard, E M; Mulieri, L A; Alpert, N R

    1990-04-03

    The heat generated by right ventricular papillary muscles of rabbits was measured after adenosine triphosphate (ATP) splitting by the contractile proteins was chemically inhibited. This tension-independent heat (TIH) (1 mJ/g wet weight) was used to calculate the total calcium (Ca) cycled in a muscle twitch by assuming that 87% of TIH was due to Ca2+ transport by the sarcoplasmic reticulum with a coupling ratio of 2 Ca2+/ATP split; the enthalpy of creatine phosphate hydrolysis buffering ATP was taken as -34 KJ/mol. The estimated Ca turnover per muscle twitch at 21 degrees C, 0.2 Hz pacing rate, and 2.5 mM Ca in the Krebs solution was approximately equal to 50 nmol/g wet weight. There was a tight positive correlation between TIH and mechanical activation during steady-state measurements but no correlation during the sharp increase in mechanical activation (treppe) when stimulation was resumed after a rest period. It is suggested that while total Ca cycling remains unchanged during the initial period of tension treppe, the free Ca2+ transient and mechanical activation increase sharply due to resaturation of high affinity Ca2+ buffers, other than troponin C, depleted of Ca2+ during the rest period.

  7. Graded boosting of synaptic signals by low-threshold voltage-activated calcium conductance

    PubMed Central

    Carbó Tano, Martín; Vilarchao, María Eugenia

    2015-01-01

    Low-threshold voltage-activated calcium conductances (LT-VACCs) play a substantial role in shaping the electrophysiological attributes of neurites. We have investigated how these conductances affect synaptic integration in a premotor nonspiking (NS) neuron of the leech nervous system. These cells exhibit an extensive neuritic tree, do not fire Na+-dependent spikes, but express an LT-VACC that was sensitive to 250 μM Ni2+ and 100 μM NNC 55-0396 (NNC). NS neurons responded to excitation of mechanosensory pressure neurons with depolarizing responses for which amplitude was a linear function of the presynaptic firing frequency. NNC decreased these synaptic responses and abolished the concomitant widespread Ca2+ signals. Coherent with the interpretation that the LT-VACC amplified signals at the postsynaptic level, this conductance also amplified the responses of NS neurons to direct injection of sinusoidal current. Synaptic amplification thus is achieved via a positive feedback in which depolarizing signals activate an LT-VACC that, in turn, boosts these signals. The wide distribution of LT-VACC could support the active propagation of depolarizing signals, turning the complex NS neuritic tree into a relatively compact electrical compartment. PMID:25972583

  8. Antibacterial activity evaluation of bioactive glass and biphasic calcium phosphate nanopowders mixtures

    NASA Astrophysics Data System (ADS)

    Nazemi, Zahra; Mehdikhani-Nahrkhalaji, Mehdi; Haghbin-Nazarpak, Masoumeh; Staji, Hamid; Kalani, Mohammad Mehdi

    2016-12-01

    The aim of this work was to evaluate the antibacterial activity of bioactive glass (BG) and biphasic calcium phosphate (BCP) nanopowders mixtures for the first time. 37S BG and BCP (50% HA-50% β-TCP) nanopowders were prepared via sol-gel technique. Characterization techniques such as X-ray diffraction, scanning electron microscopy, transition electron microscopy, and X-ray fluorescent. The antibacterial activity was studied using Escherichia coli and Salmonella typhi as gram-negative, and Staphylococcus aureus as gram-positive bacteria. The antibacterial effect of BG, BCP nanopowders, and their mixtures was evaluated at different concentrations. The 37S BG nanopowders showed minimum bactericidal concentration at 25 mg/ml. At broth concentrations below 300 mg/ml, BCP showed no antibacterial activity. BCP and BG nanopowders mixture (M2) with 60/40 ratio of BCP/BG showed noticeable antibacterial effect. It was concluded that BCP and 37S BG nanopowders mixture could be used as a good candidate for dental and orthopedic applications.

  9. Material and structural characterization of alkali activated low-calcium brown coal fly ash.

    PubMed

    Skvára, Frantisek; Kopecký, Lubomír; Smilauer, Vít; Bittnar, Zdenek

    2009-09-15

    The waste low-calcium Czech brown coal fly ash represents a considerable environmental burden due to the quantities produced and the potentially high content of leachable heavy metals. The heterogeneous microstucture of the geopolymer M(n) [-(Si-O)(z)-Al-O](n).wH(2)O, that forms during the alkaline activation, was examined by means of microcalorimetry, XRD, TGA, DSC, MIP, FTIR, NMR MAS ((29)Si, (27)Al, (23)Na), ESEM, EDS, and EBSD. The leaching of heavy metals and the evolution of compressive strength were also monitored. The analysis of raw fly ash identified a number of different morphologies, unequal distribution of elements, Fe-rich rim, high internal porosity, and minor crystalline phases of mullite and quartz. Microcalorimetry revealed exothermic reactions with dependence on the activator alkalinity. The activation energy of the geopolymerization process was determined as 86.2kJ/mol. The X-ray diffraction analysis revealed no additional crystalline phases associated with geopolymer formation. Over several weeks, the (29)Si NMR spectrum testified a high degree of polymerization and Al penetration into the SiO(4) tetrahedra. The (23)Na NMR MAS spectrum hypothesized that sodium is bound in the form of Na(H(2)O)(n) rather than Na(+), thus causing efflorescence in a moisture-gradient environment. As and Cr(6+) are weakly bonded in the geopolymer matrix, while excellent immobilization of Zn(2+), Cu(2+), Cd(2+), and Cr(3+) are reported.

  10. Effect of L-type calcium channel blocker (amlodipine) on myocardial iron deposition in patients with thalassaemia with moderate-to-severe myocardial iron deposition: protocol for a randomised, controlled trial

    PubMed Central

    Shakoor, Amarah; Zahoor, Maaman; Sadaf, Alina; Alvi, Najveen; Fadoo, Zehra; Rizvi, Arjumand; Quadri, Farheen; Tipoo, Fateh Ali; Khurshid, Mohammad; Sajjad, Zaffar; Colan, Steven; Hasan, Babar S

    2014-01-01

    Introduction Sideroblastic cardiomyopathy secondary to repeated blood transfusions is a feared complication in thalassaemia. Control of myocardial iron is thus becoming the cornerstone of thalassaemia management. Recent evidence suggests a role for L-type Ca2+ channels in mediating iron uptake by the heart. Blocking the cellular iron uptake through these channels may add to the benefit of therapy to standard chelation in reducing myocardial iron. We aim to determine the efficacy of amlodipine (a calcium channel blocker) as an adjunct to standard aggressive chelation in retarding myocardial iron deposition in thalassaemics with or without cardiomyopathy. Outcomes The primary outcome is to compare the efficacy of amlodipine+chelation (intervention) versus standard chelation (control) in retarding myocardial iron deposition. Secondary outcomes include the effect of amlodipine therapy on systolic and diastolic function, strain and strain rate and liver iron content. Methods and analysis This is a single-centre, parallel-group, prospective randomised control trial. Twenty patients will be randomised in a 1:1 allocation ratio into the intervention and control arms. In addition to conventional echocardiography, MRI T2* values for assessment of cardiac and liver iron load will be obtained at baseline and at 6 and 12 months. Cardiac T2* will be reported as the geometric mean and per cent coefficient of variation, and an increase in cardiac T2* values from baseline will be used as an end point to compare the efficacy of therapy. A p Value of <0.05 will be considered significant. Study setting Department of Pediatric and Child Health, Aga Khan University Hospital, Karachi, Pakistan. Ethics and dissemination This study has been approved by the Ethics Review Committee and Clinical Trials Unit at The Aga Khan University with respect to scientific content and compliance with applicable research and human subjects regulations. Findings will be reported through scientific

  11. Daytime deposition and nighttime dissolution of calcium carbonate controlled by submerged plants in a karst spring-fed pool: insights from high time-resolution monitoring of physico-chemistry of water

    NASA Astrophysics Data System (ADS)

    Liu, Zaihua; Liu, Xiangling; Liao, Changjun

    2008-09-01

    Water temperature, dissolved oxygen (DO), pH, and specific conductivity (spc) were measured in a time interval of 15 min in a karst spring and the spring-fed pool with flourishing submerged plants in Guilin, SW China under dry weather for periods of 2 days. Measurements allowed calculation of calcium and bicarbonate concentrations ([Ca2+] and [HCO3 -]), and thus CO2 partial pressure ( P_{{{text{CO}}2 }} ) and saturation index of calcite (SIc). Results show that there were not any diurnal variations in the physico-chemical parameters of the water for the spring. However, during daytime periods, pool water P_{{{text{CO}}2 }} decreased to far less than the spring water in a few hours, pH and SIc increased to greater than the spring, and [Ca2+] and [HCO3 -] decreased to less than the spring. During nighttime periods, pool water P_{{{text{CO}}2 }} returned to or even increased to greater than the spring, pH and SIc decreased to less than the spring, and [Ca2+] and [HCO3 -] increased to greater than the spring. The decrease in [Ca2+] and [HCO3 -] to less than the spring during daytime periods implies daytime deposition of calcium carbonate, while the increase in [Ca2+] and [HCO3 -] to greater than the spring during nighttime periods implies nighttime dissolution of calcium carbonate. The direction of the observed changes depended essentially on the illumination, indicating that daytime photosynthetic and nighttime respiratory activities in the pool aquatic plant ecosystem, which were further evidenced by the increase and decrease in DO during daytime and nighttime periods respectively, were the main processes involved. The large variations of the components of the carbonate system imply considerable changes of the capacities of CO2 and O2 in water. The finding has implications for water sampling strategy in slow-flowing karst streams and other similar environments with stagnant water bodies such as estuaries, lakes, reservoirs, and wetlands, where aquatic plant ecosystem

  12. Brain-derived neurotrophic factor inhibits calcium channel activation, exocytosis, and endocytosis at a central nerve terminal.

    PubMed

    Baydyuk, Maryna; Wu, Xin-Sheng; He, Liming; Wu, Ling-Gang

    2015-03-18

    Brain-derived neurotrophic factor (BDNF) is a neurotrophin that regulates synaptic function and plasticity and plays important roles in neuronal development, survival, and brain disorders. Despite such diverse and important roles, how BDNF, or more generally speaking, neurotrophins affect synapses, particularly nerve terminals, remains unclear. By measuring calcium currents and membrane capacitance during depolarization at a large mammalian central nerve terminal, the rat calyx of Held, we report for the first time that BDNF slows down calcium channel activation, including P/Q-type channels, and inhibits exocytosis induced by brief depolarization or single action potentials, inhibits slow and rapid endocytosis, and inhibits vesicle mobilization to the readily releasable pool. These presynaptic mechanisms may contribute to the important roles of BDNF in regulating synapses and neuronal circuits and suggest that regulation of presynaptic calcium channels, exocytosis, and endocytosis are potential mechanisms by which neurotrophins achieve diverse neuronal functions.

  13. TRPC1 regulates calcium-activated chloride channels in salivary gland cells.

    PubMed

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

    2015-11-01

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

  14. Increased calcium/calmodulin-dependent protein kinase II activity by morphine-sensitization in rat hippocampus.

    PubMed

    Kadivar, Mehdi; Farahmandfar, Maryam; Ranjbar, Faezeh Esmaeli; Zarrindast, Mohammad-Reza

    2014-07-01

    Repeated exposure to drugs of abuse, such as morphine, elicits a progressive enhancement of drug-induced behavioral responses, a phenomenon termed behavioral sensitization. These changes in behavior may reflect long-lasting changes in some of the important molecules involved in memory processing such as calcium/calmodulin-dependent protein kinase II (CaMKII). In the present study, we investigated the effect of morphine sensitization on mRNA expression of α and β isoforms and activity of CaMKII in the hippocampus of male rats. Animals were treated for 3 days with saline or morphine (20mg/kg) and following a washout period of 5 days, a challenge dose of morphine (5mg/kg) were administered. The results indicate that morphine administration in pre-treated animals produces behavioral sensitization, as determined by significant increase in locomotion and oral stereotypy behavior. In addition, repeated morphine treatment increased mRNA expression of both α and β isoforms of CaMKII in the hippocampus. The present study also showed that induction of morphine sensitization significantly increased both Ca2+/calmodulin-independent and Ca2+/calmodulin-dependent activities of CaMK II in the rat hippocampus. However, acute administration of morphine (5mg/kg) did not alter either α and β CaMKII mRNA expression or CaMKII activity in the hippocampus. The stimulation effects of morphine sensitization on mRNA expression and activity of CaMKII were completely abolished by administration of naloxone, 30min prior to s.c. injections of morphine (20mg/kg/day×3 days). Our data demonstrated that induction of morphine sensitization could effectively modulate the activity and the mRNA expression of CaMKII in the hippocampus and this effect of morphine was exerted by the activation of opioid receptors.

  15. Spectral-optical-electrical-thermal properties of deposited thin films of nano-sized calcium(II)-8-hydroxy-5,7-dinitroquinolate complex.

    PubMed

    Farag, A A M; Haggag, Sawsan M S; Mahmoud, Mohamed E

    2011-11-01

    Spectral-optical-electrical-thermal properties of deposited thin films of nano-sized calcium(II)-8-hydroxy-5,7-dinitroquinolate complex, Ca[((NO(2))(2)-8HQ)(2)], were explored, studied and evaluated in this work. Thin films of Ca[((NO(2))(2)-8HQ)(2)] were assembled by using a direct, simple and efficient layer-by-layer (LBL) chemical deposition technique. The optical properties of thin films were investigated by using spectrophotometric measurements of transmittance and reflectance at normal incidence in the wavelength range 200-2500 nm. The refractive index, n, and the absorption index, k, of Ca[((NO(2))(2)-8HQ)(2)] films were determined from the measured transmittance and reflectance. The real and imaginary dielectric constants were also determined. The analysis of the spectral behavior of the absorption coefficient in the intrinsic absorption region reveals a direct allowed transition with band gaps of 1.1 eV and 2.4 eV for the optical and transport energy gaps, respectively. The current-voltage characteristics of Ca[((NO(2))(2)-8HQ)(2)] showed a trap-charge limited conduction in determining the current at the intermediate and high bias regimes. Graphical representation of the current-voltage characteristics yields three distinct linear parts indicating the existence of three conduction mechanisms. Structural characterization and identification were confirmed by using Fourier transform infrared spectroscopy (FT-IR). Scanning electron microscopy (SEM) was also used to image the surface morphology of the deposited nano-sized metal complex and such study revealed a high homogeneity in surface spherical particle distribution with average particles size in the range 20-40 nm. Thermal gravimetric analysis (TGA) was also studied for [(NO(2))(2)-8HQ] and Ca[((NO(2))(2)-8HQ)(2)] to evaluate and confirm the thermal stability characteristics incorporated into the synthesized nano-sized Ca[((NO(2))(2)-8HQ)(2)] complex.

  16. Deoxycholic acid mediates non-canonical EGFR-MAPK activation through the induction of calcium signaling in colon cancer cells.

    PubMed

    Centuori, Sara M; Gomes, Cecil J; Trujillo, Jesse; Borg, Jamie; Brownlee, Joshua; Putnam, Charles W; Martinez, Jesse D

    2016-07-01

    Obesity and a western diet have been linked to high levels of bile acids and the development of colon cancer. Specifically, increased levels of the bile acid deoxycholic acid (DCA), an established tumor promoter, has been shown to correlate with increased development of colorectal adenomas and progression to carcinoma. Herein we investigate the mechanism by which DCA leads to EGFR-MAPK activation, a candidate mechanism by which DCA may promote colorectal tumorigenesis. DCA treated colon cancer cells exhibited strong and prolonged activation of ERK1/2 when compared to EGF treatment alone. We also showed that DCA treatment prevents EGFR degradation as opposed to the canonical EGFR recycling observed with EGF treatment. Moreover, the combination of DCA and EGF treatment displayed synergistic activity, suggesting DCA activates MAPK signaling in a non-canonical manner. Further evaluation showed that DCA treatment increased intracellular calcium levels and CAMKII phosphorylation, and that blocking calcium with BAPTA-AM abrogated MAPK activation induced by DCA, but not by EGF. Finally we showed that DCA-induced CAMKII leads to MAPK activation through the recruitment of c-Src. Taken together, we demonstrated that DCA regulates MAPK activation through calcium signaling, an alternative mechanism not previously recognized in human colon cancer cells. Importantly, this mechanism allows for EGFR to escape degradation and thus achieve a constitutively active state, which may explain its tumor promoting effects.

  17. Amyloid β Peptide Enhances RANKL-Induced Osteoclast Activation through NF-κB, ERK, and Calcium Oscillation Signaling

    PubMed Central

    Li, Shangfu; Yang, Bu; Teguh, Dian; Zhou, Lin; Xu, Jiake; Rong, Limin

    2016-01-01

    Osteoporosis and Alzheimer’s disease (AD) are common chronic degenerative disorders which are strongly associated with advanced age. We have previously demonstrated that amyloid beta peptide (Aβ), one of the pathological hallmarks of AD, accumulated abnormally in osteoporotic bone specimens in addition to having an activation effect on osteoclast (Bone 2014,61:164-75). However, the underlying molecular mechanisms remain unclear. Activation of NF-κB, extracellular signal-regulated kinase (ERK) phosphorylates, and calcium oscillation signaling pathways by receptor activator NF-κB ligand (RANKL) plays a pivotal role in osteoclast activation. Targeting this signaling to modulate osteoclast function has been a promising strategy for osteoclast-related diseases. In this study, we investigated the effects of Aβ on RANKL-induced osteoclast signaling pathways in vitro. In mouse bone marrow monocytes (BMMs), Aβ exerted no effect on RANKL-induced osteoclastogenesis but promoted osteoclastic bone resorption. In molecular levels, Aβ enhanced NF-κB activity and IκB-α degradation, activated ERK phosphorylation and stimulated calcium oscillation, thus leading to upregulation of NFAT-c1 expression during osteoclast activation. Taken together, our data demonstrate that Aβ enhances RANKL-induced osteoclast activation through IκB-α degradation, ERK phosphorylation, and calcium oscillation signaling pathways and that Aβ may be a promising agent in the treatment of osteoclast-related disease such as osteoporosis. PMID:27735865

  18. Effect of increased calcium intake on cardiac and vascular Na(+)-K(+)-ATPase activity in oral contraceptive-treated female Sprague-Dawley rats.

    PubMed

    Olatunji, L A; Soladoye, A O

    2006-11-01

    The present study aimed at investigating the influence of increased dietary calcium on Na(+)-K(+)-ATPase activity in heart and aorta of female Sprague-Dawley rats treated with oral contraceptive (OC) steroids. Rats were grouped as control (CR), OC-treated and OC+calcium-treated. OC-treated and OC+calcium-treated received a combination of OC steriods (ethinyloestradiol and norgestrel; ig). OC+calcium-treated rats were fed with 2.5% calcium diet, while OC-treated and CR groups were fed on 0.9% calcium diet. The activity of Na(+)-K(+)-ATPase in heart and aorta was significantly lower in OC-treated rats than those in the other groups. OC treatment caused significant increase in plasma glucose and significant decrease in plasma K+ as compared to control group. Decrease in Na(+)-K(+)-ATPase activity and plasma K+ was abrogated by increased calcium intake, while increase in plasma glucose was not normalized by calcium supplementation. Plasma levels of Na+, lipid peroxidation index and ascorbic acid were comparable among the three groups. These results showed that OC treatment could lead to impaired activity of cardiac and vascular Na(+)-K(+)-ATPase, possibly due to reduced plasma K+ level and these effects could be abolished by high calcium diet.

  19. CNTF-Treated Astrocyte Conditioned Medium Enhances Large-Conductance Calcium-Activated Potassium Channel Activity in Rat Cortical Neurons.

    PubMed

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

    2016-08-01

    Seizure activity is linked to astrocyte activation as well as dysfunctional cortical neuron excitability produced from changes in calcium-activated potassium (KCa) channel function. Ciliary neurotrophic factor-treated astrocyte conditioned medium (CNTF-ACM) can be used to investigate the peripheral effects of activated astrocytes upon cortical neurons. However, CNTF-ACM's effect upon KCa channel activity in cultured cortical neurons has not yet been investigated. Whole-cell patch clamp recordings were performed in rat cortical neurons to evaluate CNTF-ACM's effects upon charybdotoxin-sensitive large-conductance KCa (BK) channel currents and apamin-sensitive small-conductance KCa (SK) channel current. Biotinylation and RT-PCR were applied to assess CNTF-ACM's effects upon the protein and mRNA expression, respectively, of the SK channel subunits SK2 and SK3 and the BK channel subunits BKα1 and BKβ3. An anti-fibroblast growth factor-2 (FGF-2) monoclonal neutralizing antibody was used to assess the effects of the FGF-2 component of CNTF-ACM. CNTF-ACM significantly increased KCa channel current density, which was predominantly attributable to gains in BK channel activity (p < 0.05). CNTF-ACM produced a significant increase in BKα1 and BKβ3 expression (p < 0.05) but had no significant effect upon SK2 or SK3 expression (p > 0.05). Blocking FGF-2 produced significant reductions in KCa channel current density (p > 0.05) as well as BKα1 and BKβ3 expression in CNTF-ACM-treated neurons (p > 0.05). CNTF-ACM significantly enhances BK channel activity in rat cortical neurons and that FGF-2 is partially responsible for these effects. CNTF-induced astrocyte activation results in secretion of neuroactive factors which may affect neuronal excitability and resultant seizure activity in mammalian cortical neurons.

  20. Single-channel properties of BK-type calcium-activated potassium channels at a cholinergic presynaptic nerve terminal

    PubMed Central

    Sun, Xiao-Ping; Schlichter, Lyanne C; Stanley, Elis F

    1999-01-01

    A high-conductance calcium-activated potassium channel (BK KCa) was characterized at a cholinergic presynaptic nerve terminal using the calyx synapse isolated from the chick ciliary ganglion.The channel had a conductance of 210 pS in a 150 mM:150 mM K+ gradient, was highly selective for K+ over Na+, and was sensitive to block by external charybdotoxin or tetraethylammonium (TEA) and by internal Ba2+. At +60 mV it was activated by cytoplasmic calcium [Ca2+]i with a Kd of ≈0.5 μM and a Hill coefficient of ≈2.0. At 10 μM [Ca2+]i the channel was 50 % activated (V½) at -8.0 mV with a voltage dependence (Boltzmann slope-factor) of 32.7 mV. The V½ values hyperpolarized with an increase in [Ca2+]i while the slope factors decreased. There were no overt differences in conductance or [Ca2+]i sensitivity between BK channels from the transmitter release face and the non-release face.Open and closed times were fitted by two and three exponentials, respectively. The slow time constants were strongly affected by both [Ca2+]i and membrane potential changes.In cell-attached patch recordings BK channel opening was enhanced by a prepulse permissive for calcium influx through the patch, suggesting that the channel can be activated by calcium ion influx through neighbouring calcium channels.The properties of the presynaptic BK channel are well suited for rapid activation during the presynaptic depolarization and Ca2+ influx that are associated with transmitter release. This channel may play an important role in terminating release by rapid repolarization of the action potential. PMID:10420003

  1. Magnolol and honokiol regulate the calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli-induced diarrhea mice.

    PubMed

    Deng, Yanli; Han, Xuefeng; Tang, Shaoxun; Xiao, Wenjun; Tan, Zhiliang; Zhou, Chuanshe; Wang, Min; Kang, Jinghe

    2015-05-15

    To explore the regulatory mechanisms of magnolol and honokiol on calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli (ETEC)-induced diarrhea mice, the concentrations of serum chloride ion (Cl(-)), sodium ion (Na(+)), potassium ion (K(+)) and calcium ion (Ca(2+)) were measured. Additionally, the mRNA expressions of calmodulin 1 (CaM), calcium/calmodulin-dependent protein kinase II alpha subunit (CaMKIIα) and beta subunit (CaMKIIβ), ryanodine receptor 1, inositol 1,4,5-trisphosphate receptors (IP3 receptors), protein kinases C (PKC), potassium intermediate/small conductance calcium-activated channels (SK) and potassium large conductance calcium-activated channels(BK)were determined. A diarrhea mouse model was established using ETEC suspensions (3.29×10(9)CFU/ml) at a dosage of 0.02ml/g live body weight (BW). Magnolol or honokiol was intragastrically administered at dosages of 100 (M100 or H100), 300 (M300 or H300) and 500 (M500 or H500) mg/kg BW according to a 3×3 factorial arrangement. Magnolol and honokiol increased the Cl(-) and K(+) concentrations, further, upregulated the CaM, BKα1 and BKβ3 mRNA levels but downregulated the IP3 receptors 1, PKC, SK1, SK2, SK3, SK4 and BKβ4 mRNA expressions. Magnolol and honokiol did not alter the CaMKIIα, CaMKIIβ, ryanodine receptor 1, IP3 receptor 2, IP3 receptor 3, BKβ1 and BKβ2 mRNA expressions. These results clarify that magnolol and honokiol, acting through Ca(2+) channel blockade, inhibit the activation of IP3 receptor 1 to regulate the IP3-Ca(2+) store release, activate CaM to inhibit SK channels, and effectively suppress PKC kinases to promote BKα1 and BKβ3 channels opening and BKβ4 channel closing, which modulates the intestinal ion secretion.

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

    PubMed

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

    2014-04-01

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

  3. Enhanced calcium cycling and contractile function in transgenic hearts expressing constitutively active G alpha o* protein.

    PubMed

    Zhu, Ming; Gach, Agnieszka A; Liu, GongXin; Xu, Xiaomei; Lim, Chee Chew; Zhang, Julie X; Mao, Lan; Chuprun, Kurt; Koch, Walter J; Liao, Ronglih; Koren, Gideon; Blaxall, Burns C; Mende, Ulrike

    2008-03-01

    In contrast to the other heterotrimeric GTP-binding proteins (G proteins) Gs and Gi, the functional role of G o is still poorly defined. To investigate the role of G alpha o in the heart, we generated transgenic mice with cardiac-specific expression of a constitutively active form of G alpha o1* (G alpha o*), the predominant G alpha o isoform in the heart. G alpha o expression was increased 3- to 15-fold in mice from 5 independent lines, all of which had a normal life span and no gross cardiac morphological abnormalities. We demonstrate enhanced contractile function in G alpha o* transgenic mice in vivo, along with increased L-type Ca2+ channel current density, calcium transients, and cell shortening in ventricular G alpha o*-expressing myocytes compared with wild-type controls. These changes were evident at baseline and maintained after isoproterenol stimulation. Expression levels of all major Ca2+ handling proteins were largely unchanged, except for a modest reduction in Na+/Ca2+ exchanger in transgenic ventricles. In contrast, phosphorylation of the ryanodine receptor and phospholamban at known PKA sites was increased 1.6- and 1.9-fold, respectively, in G alpha o* ventricles. Density and affinity of beta-adrenoceptors, cAMP levels, and PKA activity were comparable in G alpha o* and wild-type myocytes, but protein phosphatase 1 activity was reduced upon G alpha o* expression, particularly in the vicinity of the ryanodine receptor. We conclude that G alpha o* exerts a positive effect on Ca2+ cycling and contractile function. Alterations in protein phosphatase 1 activity rather than PKA-mediated phosphorylation might be involved in hyperphosphorylation of key Ca2+ handling proteins in hearts with constitutive G alpha o activation.

  4. Mouse spermatozoa contain a nuclease that is activated by pretreatment with EGTA and subsequent calcium incubation.

    PubMed

    Boaz, Segal M; Dominguez, Kenneth; Shaman, Jeffrey A; Ward, W Steven

    2008-04-01

    We demonstrated that mouse spermatozoa cleave their DNA into approximately 50 kb loop-sized fragments with topoisomerase IIB when treated with MnCl(2) and CaCl(2) in a process we term sperm chromatin fragmentation (SCF). SCF can be reversed by EDTA. A nuclease then further degrades the DNA in a process we term sperm DNA degradation (SDD). MnCl(2) alone could elicit this activity, but CaCl(2) had no effect. Here, we demonstrate the existence of a nuclease in the vas deferens that can be activated by ethylene glycol tetraacetic acid (EGTA) to digest the sperm DNA by SDD. Spermatozoa were extracted with salt and dithiothreitol to remove protamines and then incubated with EGTA. Next, the EGTA was removed and divalent cations were added. We found that Mn(2+), Ca(2+), or Zn(2+) could each activate SDD in spermatozoa but Mg(2+) could not. When the reaction was slowed by incubation on ice, EGTA pretreatment followed by incubation in Ca(2+) elicited the reversible fragmentation of sperm DNA evident in SCF. When the reactions were then incubated at 37 degrees C they progressed to the more complete degradation of DNA by SDD. EDTA could also be used to activate the nuclease, but required a higher concentration than EGTA. This EGTA-activatable nuclease activity was found in each fraction of the vas deferens plasma: in the spermatozoa, in the surrounding fluid, and in the insoluble components in the fluid. These results suggest that this sperm nuclease is regulated by a mechanism that is sensitive to EGTA, possibly by removing inhibition of a calcium binding protein.

  5. Downregulation of Purkinje Cell Activity by Modulators of Small Conductance Calcium-Activated Potassium Channels In Rat Cerebellum

    PubMed Central

    Karelina, T. V.; Stepanenko, Yu. D.; Abushik, P. A.; Sibarov, D. A.; Antonov, S. M.

    2016-01-01

    Small-conductance calcium-activated potassium channels (SK channels) are widely expressed in CNS tissues. Their functions, however, have not been well studied. Participation of SK channels in Purkinje cell (PC) pacemaker activity has been studied predominantly in vitro. Here we studied for the first time the effects of SK channel activation by NS309 or CyPPA on the PC simple spike frequency in vivo in adult (3 – 6 months) and aged (22 – 28 months) rats using extracellular microelectrode recordings. Both pharmacological agents caused a statistically significant decrease in the PC simple spike frequency. The maximum value of the decrease in the simple spike frequency did not depend on age, whereas a statistically significant inhibition of the spike frequency was achieved faster in aged animals than in adult ones. In experiments on cultured neurons PCs were identified by the expression of calbindin as the PC-specific marker. Registration of transmembrane currents in cerebellar neurons revealed the direct action of NS309 and CyPPA on the SK channels of PC consisted in the enhancement of outward potassium currents and action potential after-hyperpolarization. Thus, SK channel activators can compensate for age-related changes of the autorhythmic functions of the cerebellum. PMID:28050270

  6. In vitro evaluation of antimicrobial activity of different Gutta-percha points and calcium hydroxide pastes

    PubMed Central

    Jhamb, Ashu; Chaurasia, Vishwajit Rampratap; Masamatti, Vinay kumar S.; Agarwal, Jai Hans; Tiwari, Samarth; Nair, Divya

    2014-01-01

    Purpose: To evaluate the antimicrobial activity of different compositions of Gutta-percha points and calcium hydroxide (Ca(OH)2) pastes, used in endodontic therapy. Materials and Methods: The evaluated material consisted of Gutta-percha points containing Ca(OH)2, Gutta-percha points containing chlorhexidine (Chx), conventional Gutta-percha points and Ca(OH)2 pastes. Antimicrobial properties of Chx and CaOH paste are compared with CaOH points. Antimicrobial tests included three species of microorganisms: Escherichia coli (ATCC 25923), Staphylococcus aureus (ATCC 25922) Pseudomonas aeruginosa (ATCC BAA-427), the agar disc diffusion method was employed. The plates were kept at room temperature for 2 h for prediffusion and then incubated at 37°C for 24 h. Zones of inhibition were measured. Results and Conclusion: All microbial species used in the study were inhibited by the Gutta-percha points containing Chx and by the Ca(OH)2 pastes, no antimicrobial activity was observed for the other groups (conventional Gutta-percha and Ca(OH)2 group). PMID:25254192

  7. Optimization of a GCaMP calcium indicator for neural activity imaging.

    PubMed

    Akerboom, Jasper; Chen, Tsai-Wen; Wardill, Trevor J; Tian, Lin; Marvin, Jonathan S; Mutlu, Sevinç; Calderón, Nicole Carreras; Esposti, Federico; Borghuis, Bart G; Sun, Xiaonan Richard; Gordus, Andrew; Orger, Michael B; Portugues, Ruben; Engert, Florian; Macklin, John J; Filosa, Alessandro; Aggarwal, Aman; Kerr, Rex A; Takagi, Ryousuke; Kracun, Sebastian; Shigetomi, Eiji; Khakh, Baljit S; Baier, Herwig; Lagnado, Leon; Wang, Samuel S-H; Bargmann, Cornelia I; Kimmel, Bruce E; Jayaraman, Vivek; Svoboda, Karel; Kim, Douglas S; Schreiter, Eric R; Looger, Loren L

    2012-10-03

    Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials in short bursts in several systems in vivo. Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by severalfold, creating a family of "GCaMP5" sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2- to 3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3. GCaMP5 allows more sensitive detection of neural activity in vivo and may find widespread applications for cellular imaging in general.

  8. Optimization of a GCaMP calcium indicator for neural activity imaging

    PubMed Central

    Akerboom, Jasper; Chen, Tsai-Wen; Wardill, Trevor J.; Tian, Lin; Marvin, Jonathan S.; Mutlu, Sevinç; Calderón, Nicole Carreras; Esposti, Federico; Borghuis, Bart G.; Sun, Xiaonan Richard; Gordus, Andrew; Orger, Michael B.; Portugues, Ruben; Engert, Florian; Macklin, John J.; Filosa, Alessandro; Aggarwal, Aman; Kerr, Rex; Takagi, Ryousuke; Kracun, Sebastian; Shigetomi, Eiji; Khakh, Baljit S.; Baier, Herwig; Lagnado, Leon; Wang, Samuel S.-H.; Bargmann, Cornelia I.; Kimmel, Bruce E.; Jayaraman, Vivek; Svoboda, Karel; Kim, Douglas S.; Schreiter, Eric R.; Looger, Loren L.

    2012-01-01

    Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials (APs) in short bursts in several systems in vivo. Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by several-fold, creating a family of “GCaMP5” sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2–3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3. GCaMP5 allows more sensitive detection of neural activity in vivo and may find widespread applications for cellular imaging in general. PMID:23035093

  9. Study of permeation and blocker binding in TMEM16A calcium-activated chloride channels.

    PubMed

    Reyes, J P; Huanosta-Gutiérrez, A; López-Rodríguez, A; Martínez-Torres, A

    2015-01-01

    We studied the effects of mutations of positively charged amino acid residues in the pore of X. tropicalis TMEM16A calcium-activated chloride channels: K613E, K628E, K630E; R646E and R761E. The activation and deactivation kinetics were not affected, and only K613E showed a lower current density. K628E and R761E affect anion selectivity without affecting Na(+) permeation, whereas K613E, R646E and the double mutant K613E + R646E affect anion selectivity and permeability to Na(+). Furthermore, altered blockade by the chloride channel blockers anthracene-9-carboxylic acid (A-9-C), 4, 4'-Diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) and T16inh-A01 was observed. These results suggest the existence of 2 binding sites for anions within the pore at electrical distances of 0.3 and 0.5. These sites are also relevant for anion permeation and blockade.

  10. Novel 5-substituted benzyloxy-2-arylbenzofuran-3-carboxylic acids as calcium activated chloride channel inhibitors

    PubMed Central

    Kumar, Satish; Namkung, Wan; Verkman, A. S.; Sharma, Pawan K.

    2013-01-01

    Transmembrane protein 16A (TMEM16A) channels are recently discovered membrane proteins that functions as a calcium activated chloride channel (CaCC). CaCCs are major regulators of various physiological processes, such as sensory transduction, epithelial secretion, smooth muscle contraction and oocyte fertilization. Thirty novel 5-substituted benzyloxy-2-arylbenzofuran-3-carboxylic acids (B01–B30) were synthesized and evaluated for their TMEM16A inhibitory activity by using short circuit current measurements in Fischer rat thyroid (FRT) cells expressing human TMEM16A. IC50 values were calculated using YFP fluorescence plate reader assay. Final compounds, having free carboxylic group displayed significant inhibition. Eight of the novel compounds B02, B13, B21, B23, B25, B27, B28, B29 exhibit excellent CaCCs inhibition with IC50 value <6 μM, with compound B25 exhibiting the lowest IC50 value of 2.8 ± 1.3 μM. None of the tested ester analogs of final benzofuran derivatives displayed TMEM16A/CaCCs inhibition. PMID:22739085

  11. Study of permeation and blocker binding in TMEM16A calcium-activated chloride channels

    PubMed Central

    Reyes, JP; Huanosta-Gutiérrez, A; López-Rodríguez, A; Martínez-Torres, A

    2015-01-01

    We studied the effects of mutations of positively charged amino acid residues in the pore of X. tropicalis TMEM16A calcium-activated chloride channels: K613E, K628E, K630E; R646E and R761E. The activation and deactivation kinetics were not affected, and only K613E showed a lower current density. K628E and R761E affect anion selectivity without affecting Na+ permeation, whereas K613E, R646E and the double mutant K613E + R646E affect anion selectivity and permeability to Na+. Furthermore, altered blockade by the chloride channel blockers anthracene-9-carboxylic acid (A-9-C), 4, 4'-Diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) and T16inh-A01 was observed. These results suggest the existence of 2 binding sites for anions within the pore at electrical distances of 0.3 and 0.5. These sites are also relevant for anion permeation and blockade. PMID:25853341

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

    PubMed

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

    2012-09-21

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

  13. The large conductance calcium-activated K+ channel interacts with the small GTPase Rab11b

    PubMed Central

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

    2012-01-01

    The transduction of sound by the receptor or hair cells of the cochlea leads to the activation of ion channels found in the basal and lateral regions of these cells. Thus, the processing of these transduced signals to the central nervous system is tied to the regulation of baso-lateral ion channels. The large conductance calcium-activated potassium or BK channel was revealed to interact with the small GTPase, Rab11b, which is one of many Rabs found in various endosomal pathways. Immunoelectron microscopy showed the colocalization of these two proteins in receptor cells and auditory neurons. Using Chinese hamster ovary cells as a heterologous expression system, Rab11b increased or decreased BK expression, depending on the overexpression or RNAi knockdown of Rab, respectively. Additional mutation analyses, using a yeast two-hybrid assay, suggested that this GTPase moderately interacts within a region of BK exclusive of the N- or C-terminal tails. These data suggest that this small GTPase regulates BK in a slow recycling process through the endocytic compartment and to the plasmalemma. PMID:22935415

  14. The Large Conductance, Calcium-activated K+ (BK) Channel is regulated by Cysteine String Protein

    PubMed Central

    Kyle, Barry D.; Ahrendt, Eva; Braun, Andrew P.; Braun, Janice E. A.

    2013-01-01

    Large-conductance, calcium-activated-K+ (BK) channels are widely distributed throughout the nervous system, where they regulate action potential duration and firing frequency, along with presynaptic neurotransmitter release. Our recent efforts to identify chaperones that target neuronal ion channels have revealed cysteine string protein (CSPα) as a key regulator of BK channel expression and current density. CSPα is a vesicle-associated protein and mutations in CSPα cause the hereditary neurodegenerative disorder, adult-onset autosomal dominant neuronal ceroid lipofuscinosis (ANCL). CSPα null mice show 2.5 fold higher BK channel expression compared to wild type mice, which is not seen with other neuronal channels (i.e. Cav2.2, Kv1.1 and Kv1.2). Furthermore, mutations in either CSPα's J domain or cysteine string region markedly increase BK expression and current amplitude. We conclude that CSPα acts to regulate BK channel expression, and consequently CSPα-associated changes in BK activity may contribute to the pathogenesis of neurodegenerative disorders, such as ANCL. PMID:23945775

  15. Cholinergic modulation of large-conductance calcium-activated potassium channels regulates synaptic strength and spine calcium in cartwheel cells of the dorsal cochlear nucleus.

    PubMed

    He, Shan; Wang, Ya-Xian; Petralia, Ronald S; Brenowitz, Stephan D

    2014-04-09

    Acetylcholine is a neuromodulatory transmitter that controls synaptic plasticity and sensory processing in many brain regions. The dorsal cochlear nucleus (DCN) is an auditory brainstem nucleus that integrates auditory signals from the cochlea with multisensory inputs from several brainstem nuclei and receives prominent cholinergic projections. In the auditory periphery, cholinergic modulation serves a neuroprotective function, reducing cochlear output under high sound levels. However, the role of cholinergic signaling in the DCN is less understood. Here we examine postsynaptic mechanisms of cholinergic modulation at glutamatergic synapses formed by parallel fiber axons onto cartwheel cells (CWCs) in the apical DCN circuit from mouse brainstem slice using calcium (Ca) imaging combined with two-photon laser glutamate uncaging onto CWC spines. Activation of muscarinic acetylcholine receptors (mAChRs) significantly increased the amplitude of both uncaging-evoked EPSPs (uEPSPs) and spine Ca transients. Our results demonstrate that mAChRs in CWC spines act by suppressing large-conductance calcium-activated potassium (BK) channels, and this effect is mediated through the cAMP/protein kinase A signaling pathway. Blocking BK channels relieves voltage-dependent magnesium block of NMDA receptors, thereby enhancing uEPSPs and spine Ca transients. Finally, we demonstrate that mAChR activation inhibits L-type Ca channels and thus may contribute to the suppression of BK channels by mAChRs. In summary, we demonstrate a novel role for BK channels in regulating glutamatergic transmission and show that this mechanism is under modulatory control of mAChRs.

  16. Cholinergic Modulation of Large-Conductance Calcium-Activated Potassium Channels Regulates Synaptic Strength and Spine Calcium in Cartwheel Cells of the Dorsal Cochlear Nucleus

    PubMed Central

    He, Shan; Wang, Ya-Xian; Petralia, Ronald S.

    2014-01-01

    Acetylcholine is a neuromodulatory transmitter that controls synaptic plasticity and sensory processing in many brain regions. The dorsal cochlear nucleus (DCN) is an auditory brainstem nucleus that integrates auditory signals from the cochlea with multisensory inputs from several brainstem nuclei and receives prominent cholinergic projections. In the auditory periphery, cholinergic modulation serves a neuroprotective function, reducing cochlear output under high sound levels. However, the role of cholinergic signaling in the DCN is less understood. Here we examine postsynaptic mechanisms of cholinergic modulation at glutamatergic synapses formed by parallel fiber axons onto cartwheel cells (CWCs) in the apical DCN circuit from mouse brainstem slice using calcium (Ca) imaging combined with two-photon laser glutamate uncaging onto CWC spines. Activation of muscarinic acetylcholine receptors (mAChRs) significantly increased the amplitude of both uncaging-evoked EPSPs (uEPSPs) and spine Ca transients. Our results demonstrate that mAChRs in CWC spines act by suppressing large-conductance calcium-activated potassium (BK) channels, and this effect is mediated through the cAMP/protein kinase A signaling pathway. Blocking BK channels relieves voltage-dependent magnesium block of NMDA receptors, thereby enhancing uEPSPs and spine Ca transients. Finally, we demonstrate that mAChR activation inhibits L-type Ca channels and thus may contribute to the suppression of BK channels by mAChRs. In summary, we demonstrate a novel role for BK channels in regulating glutamatergic transmission and show that this mechanism is under modulatory control of mAChRs. PMID:24719104

  17. OCT intensity and phase fluctuations correlated with activity-dependent neuronal calcium dynamics in the Drosophila CNS [Invited

    PubMed Central

    Tong, Minh Q.; Hasan, Md. Monirul; Lee, Sang Soo; Haque, Md. Rezuanul; Kim, Do-Hyoung; Islam, Md. Shahidul; Adams, Michael E.; Park, B. Hyle

    2017-01-01

    Phase-resolved OCT and fluorescence microscopy were used simultaneously to examine stereotypic patterns of neural activity in the isolated Drosophila central nervous system. Both imaging modalities were focused on individually identified bursicon neurons known to be involved in a fixed action pattern initiated by ecdysis-triggering hormone. We observed clear correspondence of OCT intensity, phase fluctuations, and activity-dependent calcium-induced fluorescence. PMID:28270979

  18. Factor Xa stimulates fibroblast procollagen production, proliferation, and calcium signaling via PAR{sub 1} activation

    SciTech Connect

    Blanc-Brude, Olivier P. . E-mail: olivier.blanc-brude@larib.inserm.fr; Archer, Fabienne; Leoni, Patricia; Derian, Claudia; Bolsover, Steven; Laurent, Geoffrey J.; Chambers, Rachel C.

    2005-03-10

    Fibroblast proliferation and procollagen production are central features of tissue repair and fibrosis. In addition to its role in blood clotting, the coagulation cascade proteinase thrombin can contribute to tissue repair by stimulating fibroblasts via proteolytic activation of proteinase-activated receptor-1 (PAR{sub 1}). During hemostasis, the coagulation cascade proteinase factor X is converted into factor Xa. We have previously shown that factor Xa upregulates fibroblast proliferation via production of autocrine PDGF. In this study, we further examined the effects of factor Xa on fibroblast function and aimed to identify its signaling receptor. We showed that factor Xa stimulates procollagen promoter activity and protein production by human and mouse fibroblasts. This effect was independent of PDGF and thrombin production, but dependent on factor Xa proteolytic activity. We also showed that PAR{sub 1}-deficient mouse fibroblasts did not upregulate procollagen production, mobilize cytosolic calcium, or proliferate in response to factor Xa. Desensitization techniques and PAR{sub 1}-specific agonists and inhibitors were used to demonstrate that PAR{sub 1} mediates factor Xa signaling in human fibroblasts. This is the first report that factor Xa stimulates extracellular matrix production. In contrast with endothelial cells and vascular smooth muscle cells, fibroblasts appear to be the only cell type in which the effects of factor Xa are mediated mainly via PAR{sub 1} and not PAR{sub 2}. These findings are critical for our understanding of tissue repair and fibrotic mechanisms, and for the design of novel approaches to inhibit the profibrotic effects of the coagulation cascade without compromising blood hemostasis.

  19. Plasma Membrane Calcium ATPase Activity Is Regulated by Actin Oligomers through Direct Interaction*

    PubMed Central

    Dalghi, Marianela G.; Fernández, Marisa M.; Ferreira-Gomes, Mariela; Mangialavori, Irene C.; Malchiodi, Emilio L.; Strehler, Emanuel E.; Rossi, Juan Pablo F. C.

    2013-01-01

    As recently described by our group, plasma membrane calcium ATPase (PMCA) activity can be regulated by the actin cytoskeleton. In this study, we characterize the interaction of purified G-actin with isolated PMCA and examine the effect of G-actin during the first polymerization steps. As measured by surface plasmon resonance, G-actin directly interacts with PMCA with an apparent 1:1 stoichiometry in the presence of Ca2+ with an apparent affinity in the micromolar range. As assessed by the photoactivatable probe 1-O-hexadecanoyl-2-O-[9-[[[2-[125I]iodo-4-(trifluoromethyl-3H-diazirin-3-yl)benzyl]oxy]carbonyl]nonanoyl]-sn-glycero-3-phosphocholine, the association of PMCA to actin produced a shift in the distribution of the conformers of the pump toward a calmodulin-activated conformation. G-actin stimulates Ca2+-ATPase activity of the enzyme when incubated under polymerizing conditions, displaying a cooperative behavior. The increase in the Ca2+-ATPase activity was related to an increase in the apparent affinity for Ca2+ and an increase in the phosphoenzyme levels at steady state. Although surface plasmon resonance experiments revealed only one binding site for G-actin, results clearly indicate that more than one molecule of G-actin was needed for a regulatory effect on the pump. Polymerization studies showed that the experimental conditions are compatible with the presence of actin in the first stages of assembly. Altogether, these observations suggest that the stimulatory effect is exerted by short oligomers of actin. The functional interaction between actin oligomers and PMCA represents a novel regulatory pathway by which the cortical actin cytoskeleton participates in the regulation of cytosolic Ca2+ homeostasis. PMID:23803603

  20. Plasma membrane calcium ATPase activity is regulated by actin oligomers through direct interaction.

    PubMed

    Dalghi, Marianela G; Fernández, Marisa M; Ferreira-Gomes, Mariela; Mangialavori, Irene C; Malchiodi, Emilio L; Strehler, Emanuel E; Rossi, Juan Pablo F C

    2013-08-09

    As recently described by our group, plasma membrane calcium ATPase (PMCA) activity can be regulated by the actin cytoskeleton. In this study, we characterize the interaction of purified G-actin with isolated PMCA and examine the effect of G-actin during the first polymerization steps. As measured by surface plasmon resonance, G-actin directly interacts with PMCA with an apparent 1:1 stoichiometry in the presence of Ca(2+) with an apparent affinity in the micromolar range. As assessed by the photoactivatable probe 1-O-hexadecanoyl-2-O-[9-[[[2-[(125)I]iodo-4-(trifluoromethyl-3H-diazirin-3-yl)benzyl]oxy]carbonyl]nonanoyl]-sn-glycero-3-phosphocholine, the association of PMCA to actin produced a shift in the distribution of the conformers of the pump toward a calmodulin-activated conformation. G-actin stimulates Ca(2+)-ATPase activity of the enzyme when incubated under polymerizing conditions, displaying a cooperative behavior. The increase in the Ca(2+)-ATPase activity was related to an increase in the apparent affinity for Ca(2+) and an increase in the phosphoenzyme levels at steady state. Although surface plasmon resonance experiments revealed only one binding site for G-actin, results clearly indicate that more than one molecule of G-actin was needed for a regulatory effect on the pump. Polymerization studies showed that the experimental conditions are compatible with the presence of actin in the first stages of assembly. Altogether, these observations suggest that the stimulatory effect is exerted by short oligomers of actin. The functional interaction between actin oligomers and PMCA represents a novel regulatory pathway by which the cortical actin cytoskeleton participates in the regulation of cytosolic Ca(2+) homeostasis.

  1. Antibacterial activity of TiO2 nanotubes: Influence of crystal phase, morphology and Ag deposition

    NASA Astrophysics Data System (ADS)

    Li, Huirong; Cui, Qiang; Feng, Bo; Wang, Jianxin; Lu, Xiong; Weng, Jie

    2013-11-01

    TiO2 nanotubes on Ti substrate were fabricated by electrochemical anodization. Ag nanoparticles were deposited on the TiO2 nanotubes by a silver mirror reaction. Antibacterial activity of the nanotubes with different structural features was evaluated by a culture test with Escherichia coli bacteria. The anatase nanotubes showed the highest antibacterial activity among three crystal phases including anatase, rutile and amorphous titania. The diameters of the nanotubes affected the antibacterial activity. The two nanotubes with 200 nm and 50 nm diameters had higher antibacterial rate than those with other diameters. The antibacterial activity of the nanotubes was independent on their lengths. Ag-deposited nanotubes exhibited excellent antibacterial activity and its antibacterial rate was up to approximately 100%. TiO2 nanotubes and Ag-deposited nanotubes on titanium should be potential for antibacterial applications in clinics and industry, especially regarding with their reusability.

  2. Effect of Pd precursor status on sonochemical surface activation in Cu electroless deposition

    NASA Astrophysics Data System (ADS)

    Kim, Kanghoon; Jin, Seonok; Kwon, Oh Joong

    2016-02-01

    Pd surface activation via a sonochemical approach was studied by varying Pd precursor status in the aqueous solution. By aging a K2PdCl6 activation solution overnight with added NH4OH, the chlorinated Pd complex was changed to an ammonia-based Pd complex. The Pd surface activation carried out with the NH4OH complexing agent resulted in improved surface condition following Cu electroless deposition. The Cu thin film deposited on a substrate sonochemically activated with the aged, ammonia-based Pd complex showed improved surface roughness and resistivity compared to that for Cu films deposited via two other precursors (chlorinated Pd complex and ammonia-based complex) without aging. In addition, nitrogen purging during sonochemical activation improved Cu film quality.

  3. Calcium Transients in Dendrites of Neocortical Neurons Evoked by Single Subthreshold Excitatory Postsynaptic Potentials via Low-Voltage-Activated Calcium Channels

    NASA Astrophysics Data System (ADS)

    Markram, Henry; Sakmann, Bert

    1994-05-01

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

  4. Role of zinc influx via AMPA/kainate receptor activation in metabotropic glutamate receptor-mediated calcium release.

    PubMed

    Takeda, Atsushi; Fuke, Sayuri; Minami, Akira; Oku, Naoto

    2007-05-01

    The uptake of free zinc into CA3 pyramidal cells and its significance was examined in rat hippocampal slices with ZnAF-2DA, a membrane-permeable zinc indicator. Intracellular ZnAF-2 signal in the CA3 pyramidal cell layer was increased during delivery of tetanic stimuli to the dentate granule cell layer. This increase was completely blocked in the presence of CNQX, an AMPA/kainate receptor antagonist. These results suggest that free zinc is taken up into CA3 pyramidal cells via activation of AMPA/kainate receptors. The effect of free zinc levels in the CA3 pyramidal cells on the increase in intracellular calcium via Group I metabotropic glutamate receptors was examined by regional delivery of tADA, a Group I metabotropic glutamate receptor agonist, to the stratum lucidum after blockade of AMPA/kainate receptor-mediated calcium and zinc influx. Intracellular calcium orange signal in the CA3 pyramidal cell layer was increased by tADA, whereas intracellular ZnAF-2 signal was not increased even in the presence of 100 muM zinc, suggesting that tADA induces calcium release from internal stores in CA3 pyramidal cells and is not involved in zinc uptake. The increase in calcium orange signal by tADA was enhanced by perfusion with pyrithione, a zinc ionophore that decreased basal ZnAF-2 signal in the CA3 pyramidal cell layer. It was blocked by perfusion with pyrithione and zinc that increased basal ZnAF-2 signal. The present study indicates that the increase in free calcium levels via the metabotropic glutamate receptor pathway is inversely related to free zinc levels in CA3 pyramidal cells.

  5. Mapping calcium phosphate activated gene networks as a strategy for targeted osteoinduction of human progenitors in vitro and in vivo

    PubMed Central

    Eyckmans, J.; Roberts, S.J.; Bolander, J.; Schrooten, J.; Chen, C.S.; Luyten, F.P.

    2014-01-01

    Although calcium phosphate-containing biomaterials are promising scaffolds for bone regenerative strategies, the osteoinductive capacity of such materials is poorly understood. In this study, we investigated whether endogenous mechanisms of in vivo calcium phosphate-driven, ectopic bone formation could be identified and used to induce enhanced differentiation in vitro of the same progenitor population. To accomplish this, human periosteum derived cells (hPDCs) were seeded on hydroxyapatite/collagen scaffolds (calcium phosphate rich matrix or CPRM), or on decalcified scaffolds (calcium phosphate depleted matrix or CPDM), followed by subcutaneous implantation in nude mice to trigger ectopic bone formation. In this system, osteoblast differentiation occurred in CPRM scaffolds, but not in CPDM scaffolds. Gene expression was assessed by human full-genome microarray at 20 hours after seeding, and 2, 8 and 18 days after implantation. In both matrices, implantation of the cell constructs triggered a similar gene expression cascade, however, gene expression dynamics progressed faster in CPRM scaffolds than in CPDM scaffolds. The difference in gene expression dynamics was associated with differential activation of hub genes and molecular signaling pathways related to calcium signaling (CREB), inflammation (TNFα, NFkB, and IL6) and bone development (TGFβ, β-catenin, BMP, EGF, and ERK signaling). Starting from this set of pathways, a growth factor cocktail was developed that robustly enhanced osteogenesis in vitro and in vivo. Taken together, our data demonstrate that through the identification and subsequent stimulation of genes, proteins and signaling pathways associated with calcium phosphate mediated osteoinduction, a focused approach to develop targeted differentiation protocols in adult progenitor cells can be achieved. PMID:23537666

  6. Dexamethasone-induced cardiac deterioration is associated with both calcium handling abnormalities and calcineurin signaling pathway activation.

    PubMed

    de Salvi Guimarães, Fabiana; de Moraes, Wilson Max Almeida Monteiro; Bozi, Luis Henrique Marchesi; Souza, Pâmela R; Antonio, Ednei Luiz; Bocalini, Danilo Sales; Tucci, Paulo José Ferreira; Ribeiro, Daniel Araki; Brum, Patricia Chakur; Medeiros, Alessandra

    2017-01-01

    Dexamethasone is a potent and widely used anti-inflammatory and immunosuppressive drug. However, recent evidences suggest that dexamethasone cause pathologic cardiac remodeling, which later impairs cardiac function. The mechanism behind the cardiotoxic effect of dexamethasone is elusive. The present study aimed to verify if dexamethasone-induced cardiotoxicity would be associated with changes in the cardiac net balance of calcium handling protein and calcineurin signaling pathway activation. Wistar rats (~400 g) were treated with dexamethasone (35 µg/g) in drinking water for 15 days. After dexamethasone treatment, we analyzed cardiac function, cardiomyocyte diameter, cardiac fibrosis, and the expression of proteins involved in calcium handling and calcineurin signaling pathway. Dexamethasone-treated rats showed several cardiovascular abnormalities, including elevated blood pressure, diastolic dysfunction, cardiac fibrosis, and cardiomyocyte apoptosis. Regarding the expression of proteins involved in calcium handling, dexamethasone increased phosphorylation of phospholamban at threonine 17, reduced protein levels of Na(+)/Ca(2+) exchanger, and had no effect on protein expression of Serca2a. Protein levels of NFAT and GATA-4 were increased in both cytoplasmic and nuclear faction. In addition, dexamethasone increased nuclear protein levels of calcineurin. Altogether our findings suggest that dexamethasone causes pathologic cardiac remodeling and diastolic dysfunction, which is associated with impaired calcium handling and calcineurin signaling pathway activation.

  7. Calcium-Activated Chloride Channels (CaCCs) Regulate Action Potential and Synaptic Response in Hippocampal Neurons

    PubMed Central

    Huang, Wendy C.; Xiao, Shaohua; Huang, Fen; Harfe, Brian D.; Jan, Yuh Nung; Jan, Lily Yeh

    2012-01-01

    SUMMARY Central neurons respond to synaptic inputs from other neurons by generating synaptic potentials. Once the summated synaptic potentials reach threshold for action potential firing, the signal propagates leading to transmitter release at the synapse. The calcium influx accompanying such signaling opens calcium-activated ion channels for feedback regulation. Here we report a novel mechanism for modulating hippocampal neuronal signaling that involves calcium-activated chloride channels (CaCCs). We present the first evidence that CaCCs reside in hippocampal neurons and are in close proximity of calcium channels and NMDA receptors to shorten action potential duration, dampen excitatory synaptic potentials, impede temporal summation, and raise the threshold for action potential generation by synaptic potential. Having recently identified TMEM16A and TMEM16B as CaCCs, we further show that TMEM16B but not TMEM16A is important for hippocampal CaCC, laying the groundwork for deciphering the dynamic CaCC modulation of neuronal signaling in neurons important for learning and memory. PMID:22500639

  8. Combining modelling and experimental approaches to explain how calcium signatures are decoded by calmodulin-binding transcription activators (CAMTAs) to produce specific gene expression responses.

    PubMed

    Liu, Junli; Whalley, Helen J; Knight, Marc R

    2015-10-01

    Experimental data show that Arabidopsis thaliana is able to decode different calcium signatures to produce specific gene expression responses. It is also known that calmodulin-binding transcription activators (CAMTAs) have calmodulin (CaM)-binding domains. Therefore, the gene expression responses regulated by CAMTAs respond to calcium signals. However, little is known about how different calcium signatures are decoded by CAMTAs to produce specific gene expression responses. A dynamic model of Ca(2+) -CaM-CAMTA binding and gene expression responses is developed following thermodynamic and kinetic principles. The model is parameterized using experimental data. Then it is used to analyse how different calcium signatures are decoded by CAMTAs to produce specific gene expression responses. Modelling analysis reveals that: calcium signals in the form of cytosolic calcium concentration elevations are nonlinearly amplified by binding of Ca(2+) , CaM and CAMTAs; amplification of Ca(2+) signals enables calcium signatures to be decoded to give specific CAMTA-regulated gene expression responses; gene expression responses to a calcium signature depend upon its history and accumulate all the information during the lifetime of the calcium signature. Information flow from calcium signatures to CAMTA-regulated gene expression responses has been established by combining experimental data with mathematical modelling.

  9. Sleep- and wake-dependent changes in neuronal activity and reactivity demonstrated in fly neurons using in vivo calcium imaging.

    PubMed

    Bushey, Daniel; Tononi, Giulio; Cirelli, Chiara

    2015-04-14

    Sleep in Drosophila shares many features with mammalian sleep, but it remains unknown whether spontaneous and evoked activity of individual neurons change with the sleep/wake cycle in flies as they do in mammals. Here we used calcium imaging to assess how the Kenyon cells in the fly mushroom bodies change their activity and reactivity to stimuli during sleep, wake, and after short or long sleep deprivation. As before, sleep was defined as a period of immobility of >5 min associated with a reduced behavioral response to a stimulus. We found that calcium levels in Kenyon cells decline when flies fall asleep and increase when they wake up. Moreover, calcium transients in response to two different stimuli are larger in awake flies than in sleeping flies. The activity of Kenyon cells is also affected by sleep/wake history: in awake flies, more cells are spontaneously active and responding to stimuli if the last several hours (5-8 h) before imaging were spent awake rather than asleep. By contrast, long wake (≥29 h) reduces both baseline and evoked neural activity and decreases the ability of neurons to respond consistently to the same repeated stimulus. The latter finding may underlie some of the negative effects of sleep deprivation on cognitive performance and is consistent with the occurrence of local sleep during wake as described in behaving rats. Thus, calcium imaging uncovers new similarities between fly and mammalian sleep: fly neurons are more active and reactive in wake than in sleep, and their activity tracks sleep/wake history.

  10. 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 Excess1[C][W][OA

    PubMed Central

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

    2012-01-01

    To analyze the copper-induced cross talk among calcium, nitric oxide (NO), and hydrogen peroxide (H2O2) 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 H2O2, 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 H2O2 increased at 2, 3, and 12 h and was inhibited by ned-19, ryanodine, xestospongin C, and moniliformin, indicating that H2O2 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 H2O2. 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, H2O2, and NO and a calcium-dependent activation of gene expression involving calmodulins and calcium-dependent protein kinases. PMID:22234999

  11. 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.

    PubMed

    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

  12. Zolpidem reduces hippocampal neuronal activity in freely behaving mice: a large scale calcium imaging study with miniaturized fluorescence microscope.

    PubMed

    Berdyyeva, Tamara; Otte, Stephani; Aluisio, Leah; Ziv, Yaniv; Burns, Laurie D; Dugovic, Christine; Yun, Sujin; Ghosh, Kunal K; Schnitzer, Mark J; Lovenberg, Timothy; Bonaventure, Pascal

    2014-01-01

    Therapeutic drugs for cognitive and psychiatric disorders are often characterized by their molecular mechanism of action. Here we demonstrate a new approach to elucidate drug action on large-scale neuronal activity by tracking somatic calcium dynamics in hundreds of CA1 hippocampal neurons of pharmacologically manipulated behaving mice. We used an adeno-associated viral vector to express the calcium sensor GCaMP3 in CA1 pyramidal cells under control of the CaMKII promoter and a miniaturized microscope to observe cellular dynamics. We visualized these dynamics with and without a systemic administration of Zolpidem, a GABAA agonist that is the most commonly prescribed drug for the treatment of insomnia in the United States. Despite growing concerns about the potential adverse effects of Zolpidem on memory and cognition, it remained unclear whether Zolpidem alters neuronal activity in the hippocampus, a brain area critical for cognition and memory. Zolpidem, when delivered at a dose known to induce and prolong sleep, strongly suppressed CA1 calcium signaling. The rate of calcium transients after Zolpidem administration was significantly lower compared to vehicle treatment. To factor out the contribution of changes in locomotor or physiological conditions following Zolpidem treatment, we compared the cellular activity across comparable epochs matched by locomotor and physiological assessments. This analysis revealed significantly depressive effects of Zolpidem regardless of the animal's state. Individual hippocampal CA1 pyramidal cells differed in their responses to Zolpidem with the majority (∼ 65%) significantly decreasing the rate of calcium transients, and a small subset (3%) showing an unexpected and significant increase. By linking molecular mechanisms with the dynamics of neural circuitry and behavioral states, this approach has the potential to contribute substantially to the development of new therapeutics for the treatment of CNS disorders.

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

    PubMed

    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 < 0.05), (ii) elevation in ΔΨm (p < 0.05), (iii) increased OCR and ATP formation (p < 0.05), (iv) increased intracellular NO levels (p < 0.05), (v) increased mitochondrial ROS production (p < 0.05), and (vi) increased susceptibility to rotenone (p < 0.05). Treatment with isradipine was able to partially rescue these negative effects of CNTF-ACM (p < 0.05). CNTF-ACM promotes mitochondrial respiration and oxidative stress in cortical neurons through elevating L-type calcium channel activity.

  14. Chlorotoxin does not inhibit volume-regulated, calcium-activated and cyclic AMP-activated chloride channels

    PubMed Central

    Maertens, Chantal; Wei, Lin; Tytgat, Jan; Droogmans, Guy; Nilius, Bernd

    2000-01-01

    It was the aim of this study to look for a high-affinity and selective polypeptide toxin, which could serve as a probe for the volume-regulated anion channel (VRAC) or the calcium-activated chloride channel (CaCC). We have partially purified chlorotoxin, including new and homologous short chain insectotoxins, from the crude venom of Leiurus quinquestriatus quinquestriatus (Lqq) by means of gel filtration chromatography. Material eluting between 280 and 420 min, corresponding to fractions 15–21, was lyophilized and tested on VRAC and CaCC, using the whole-cell patch-clamp technique. We have also tested the commercially available chlorotoxin on VRAC, CaCC, the cystic fibrosis transmembrane conductance regulator (CFTR) and on the glioma specific chloride channel (GCC). VRAC and the correspondent current, ICl,swell, was activated in Cultured Pulmonary Artery Endothelial (CPAE) cells by a 25% hypotonic solution. Neither of the fractions 16–21 significantly inhibited ICl,swell (n=4–5). Ca2+-activated Cl− currents, ICl,Ca, activated by loading T84 cells via the patch pipette with 1 μM free Ca2+, were not inhibited by any of the tested fractions (15–21), (n=2–5). Chlorotoxin (625 nM) did neither effect ICl,swell nor ICl,Ca (n=4–5). The CFTR channel, transiently transfected in COS cells and activated by a cocktail containing IBMX and forskolin, was not affected by 1.2 μM chlorotoxin (n=5). In addition, it did not affect currents through GCC. We conclude that submicromolar concentrations of chlorotoxin do not block volume-regulated, Ca2+-activated and CFTR chloride channels and that it can not be classified as a general chloride channel toxin. PMID:10683204

  15. Endogenous intracellular calcium buffering and the activation/inactivation of HVA calcium currents in rat dentate gyrus granule cells

    PubMed Central

    1991-01-01

    Granule cells acutely dissociated from the dentate gyrus of adult rat brains displayed a single class of high-threshold, voltage-activated (HVA) Ca2+ channels. The kinetics of whole-cell Ca2+ currents recorded with pipette solutions containing an intracellular ATP regenerating system but devoid of exogenous Ca2+ buffers, were fit best by Hodgkin- Huxley kinetics (m2h), and were indistinguishable from those recorded with the nystatin perforated patch method. In the absence of exogenous Ca2+ buffers, inactivation of HVA Ca2+ channels was a predominantly Ca(2+)-dependent process. The contribution of endogenous Ca2+ buffers to the kinetics of inactivation was investigated by comparing currents recorded from control cells to currents recorded from neurons that have lost a specific Ca(2+)-binding protein, Calbindin-D28K (CaBP), after kindling-induced epilepsy. Kindled neurons devoid of CaBP showed faster rates of both activation and inactivation. Adding an exogenous Ca2+ chelator, 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), to the intracellular solution largely eliminated inactivation in both control and kindled neurons. The results are consistent with the hypothesis that endogenous intraneuronal CaBP contributes significantly to submembrane Ca2+ sequestration at a concentration range and time domain that regulate Ca2+ channel inactivation. PMID:1662686

  16. Calcium-calmodulin does not alter the anion permeability of the mouse TMEM16A calcium-activated chloride channel

    PubMed Central

    Yu, Yawei; Kuan, Ai-Seon

    2014-01-01

    The transmembrane protein TMEM16A forms a Ca2+-activated Cl− channel that is permeable to many anions, including SCN−, I−, Br−, Cl−, and HCO3−, and has been implicated in various physiological functions. Indeed, controlling anion permeation through the TMEM16A channel pore may be critical in regulating the pH of exocrine fluids such as the pancreatic juice. The anion permeability of the TMEM16A channel pore has recently been reported to be modulated by Ca2+-calmodulin (CaCaM), such that the pore of the CaCaM-bound channel shows a reduced ability to discriminate between anions as measured by a shift of the reversal potential under bi-ionic conditions. Here, using a mouse TMEM16A clone that contains the two previously identified putative CaM-binding motifs, we were unable to demonstrate such CaCaM-dependent changes in the bi-ionic potential. We confirmed the activity of CaCaM used in our study by showing CaCaM modulation of the olfactory cyclic nucleotide–gated channel. We suspect that the different bi-ionic potentials that were obtained previously from whole-cell recordings in low and high intracellular [Ca2+] may result from different degrees of bi-ionic potential shift secondary to a series resistance problem, an ion accumulation effect, or both. PMID:24981232

  17. The structure-activity relationship study on 2-, 5-, and 6-position of the water soluble 1,4-dihydropyridine derivatives blocking N-type calcium channels.

    PubMed

    Yamamoto, Takashi; Niwa, Seiji; Ohno, Seiji; Tokumasu, Munetaka; Masuzawa, Yoko; Nakanishi, Chika; Nakajo, Akira; Onishi, Tomoyuki; Koganei, Hajime; Fujita, Shin-Ichi; Takeda, Tomoko; Kito, Morikazu; Ono, Yukitsugu; Saitou, Yuki; Takahara, Akira; Iwata, Seinosuke; Shoji, Masataka

    2008-09-01

    In order to find an injectable and selective N-type calcium channel blocker, we have performed the structure-activity relationship (SAR) study on the 2-, 5-, and 6-position of 1,4-dihydropyridine-3-carboxylate derivative APJ2708 (2), which is a derivative of Cilnidipine and has L/N-type calcium channel dual inhibitory activities. As a consequence of the optimization, 6-dimethylacetal derivative 7 was found to have an effective inhibitory activity against N-type calcium channels with more than 170-fold lower activity for L-type channel compared to that of APJ2708.

  18. Identifying calcium sources at an acid deposition-impacted spruce forest: A strontium isotope, alkaline earth element multi-tracer approach

    USGS Publications Warehouse

    Bullen, T.D.; Bailey, S.W.

    2005-01-01

    Depletion of calcium from forest soils has important implications for forest productivity and health. Ca is available to fine feeder roots from a number of soil organic and mineral sources, but identifying the primary source or changes of sources in response to environmental change is problematic. We used strontium isotope and alkaline earth element concentration ratios of trees and soils to discern the record of Ca sources for red spruce at a base-poor, acid deposition-impacted watershed. We measured 87Sr/86Sr and chemical compositions of cross-sectional stemwood cores of red spruce, other spruce tissues and sequential extracts of co-located soil samples. 87Sr/86Sr and Sr/Ba ratios together provide a tracer of alkaline earth element sources that distinguishes the plant-available fraction of the shallow organic soils from those of deeper organic and mineral soils. Ca/Sr ratios proved less diagnostic, due to within-tree processes that fractionate these elements from each other. Over the growth period from 1870 to 1960, 87Sr/86Sr and Sr/Ba ratios of stemwood samples became progressively more variable and on average trended toward values that considered together are characteristic of the uppermost forest floor. In detail the stemwood chemistry revealed an episode of simultaneous enhanced uptake of all alkaline earth elements during the growth period from 1930 to 1960, coincident with reported local and regional increases in atmospheric inputs of inorganic acidity. We attribute the temporal trends in stemwood chemistry to progressive shallowing of the effective depth of alkaline earth element uptake by fine roots over this growth period, due to preferential concentration of fine roots in the upper forest floor coupled with reduced nutrient uptake by roots in the lower organic and upper mineral soils in response to acid-induced aluminum toxicity. Although both increased atmospheric deposition and selective weathering of Ca-rich minerals such as apatite provide possible

  19. Simultaneous monitoring of biofilm growth, microbial activity, and inorganic deposits on surfaces with an in situ, online, real-time, non-destructive, optical sensor.

    PubMed

    Strathmann, Martin; Mittenzwey, Klaus-Henrik; Sinn, Gert; Papadakis, Wassilios; Flemming, Hans-Curt

    2013-01-01

    Deposits on surfaces in water-bearing systems, also known as 'fouling', can lead to substantial losses in the performance of industrial processes as well as a decreased product quality. Early detection and localization of such deposits can, to a considerable extent, save such losses. However, most of the surfaces that become fouled, for example, in process water pipes, membrane systems, power plants, and food and beverage industries, are difficult to access and analyses conducted on the water phase do not reveal the site or extent of deposits. Furthermore, it is of interest to distinguish biological from non-biological deposits. Although they usually occur together, different countermeasures are necessary. Therefore, sensors are required that indicate the development of surface fouling in real-time, non-destructively, and in situ, preferably allowing for discrimination between chemical and/or biological deposits. In this paper, an optical deposit sensor is presented which fulfills these requirements. Based on multiple fluorescence excitation emission matrix analysis, it detects autofluorescence of amino acids as indicators of biomass. Autofluorescence of nicotinamide adenine dinucleotide + hydrogen is interpreted as an indicator of biological activity, thus it acts as a viability marker, making the method suited for assessing the efficacy of disinfection treatments. Scattering signals from abiotic deposits such as calcium carbonate or corrosion products can clearly be distinguished from biotic substances and monitored separately. The sensor provides an early warning of fouling, allowing for timely countermeasures to be deployed. It also provides an assessment of the success of cleaning treatments and is a promising tool for integrated antifouling strategies.

  20. A 125 year record of fluvial calcium flux from a temperate catchment: Interplay of climate, land-use change and atmospheric deposition