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Cytosolic Ca2+ Concentration (cytosolic + ca2+_concentration)
Selected AbstractsTrimetazidine Reduces Basal Cytosolic Ca2+ Concentration During Hypoxia in Single Xenopus Skeletal MyocytesEXPERIMENTAL PHYSIOLOGY, Issue 3 2003C. M. Stary We tested the hypotheses that: (1) Ca2+ handling and force production would be irreversibly altered in skeletal muscle during steady-state contractions when subjected to severe, prolonged hypoxia and subsequent reoxygenation; and (2) application of the cardio-protective drug trimetazidine would attenuate these alterations. Single, living skeletal muscle fibres from Xenopus laevis were injected with the Ca2+ indicator fura 2, and incubated for 1 h prior to stimulation in 100 ,M TMZ-Ringer solution (TMZ; n = 6) or standard Ringer solution (CON; n = 6). Force and relative free cytosolic Ca2+ concentration ([Ca2+]c) were measured during continuous tetanic contractions produced every 5 s as fibres were sequentially perfused in the following manner: 3 min high extracellular PO2 (159 mmHg), 15 min hypoxic perfusion (3-5 mmHg) then 3 min high PO2. Hypoxia caused a decrease in force and peak [Ca2+]c in both the TMZ and CON fibres, with no significant (P < 0.05) difference between groups. However, basal [Ca2+]c was significantly lower during hypoxia in the TMZ group vs. the CON group. While reoxygenation generated only modest recovery of relative force and peak [Ca2+]c in both groups, basal [Ca2+]c remained significantly less in the TMZ group. These results demonstrated that in contracting, single skeletal muscle fibres, TMZ prevented increases in basal [Ca2+]c generated during a severe hypoxic insult and subsequent reoxygenation, yet failed to protect the cell from the deleterious effects of prolonged hypoxia followed by reoxygenation. [source] Cytosolic Ca2+ concentration and rate of increase of the cytosolic Ca2+ concentration in the regulation of vascular permeability in Rana in vivoTHE JOURNAL OF PHYSIOLOGY, Issue 3 2005C. A. Glass Vascular permeability is assumed to be regulated by the cytosolic Ca2+ concentration ([Ca2+]c) of the endothelial cells. When permeability is increased, however, the maximum [Ca2+]c appears to occur after the maximum permeability increase, suggesting that Ca2+ -dependent mechanisms other than the absolute Ca2+ concentration may regulate permeability. Here we investigate whether the rate of increase of the [Ca2+]c (d[Ca2+]c/dt) may more closely approximate the time course of the permeability increase. Hydraulic conductivity (Lp) and endothelial [Ca2+]c were measured in single perfused frog mesenteric microvessels in vivo. The relationships between the time courses of the increased Lp, [Ca2+]c and d[Ca2+]c/dt were examined. Lp peaked significantly earlier than [Ca2+]c in all drug treatments examined (Ca2+ store release, store-mediated Ca2+ influx, and store-independent Ca2+ influx). When Lp was increased in a store-dependent manner the time taken for Lp to peak (3.6 ± 0.9 min during store release, 1.2 ± 0.3 min during store-mediated Ca2+ influx) was significantly less than the time taken for [Ca2+]c to peak (9.2 ± 2.8 min during store release, 2.1 ± 0.7 min during store-mediated influx), but very similar to that for the peak d[Ca2+]c/dt to occur (4.3 ± 2.0 min during store release, 1.1 ± 0.5 min during Ca2+ influx). Additionally, when the increase was independent of intracellular Ca2+ stores, Lp (0.38 ± 0.03 min) and d[Ca2+]c/dt (0.30 ± 0.1 min) both peaked significantly before the [Ca2+]c (1.05 ± 0.31 min). These data suggest that the regulation of vascular permeability by endothelial cell Ca2+ may be regulated by the rate of change of the [Ca2+]c rather than the global [Ca2+]. [source] Pre- and postsynaptic contributions of voltage-dependent Ca2+ channels to nociceptive transmission in rat spinal lamina I neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2004B. Heinke Abstract Activation of voltage-dependent Ca2+ channels (VDCCs) is critical for neurotransmitter release, neuronal excitability and postsynaptic Ca2+ signalling. Antagonists of VDCCs can be antinociceptive in different animal pain models. Neurons in lamina I of the spinal dorsal horn play a pivotal role in the processing of pain-related information, but the role of VDCCs to the activity-dependent Ca2+ increase in lamina I neurons and to the synaptic transmission between nociceptive afferents and second order neurons in lamina I is not known. This has now been investigated in a lumbar spinal cord slice preparation from young Sprague,Dawley rats. Microfluorometric Ca2+ measurements with fura-2 have been used to analyse the Ca2+ increase in lamina I neurons after depolarization of the cells, resulting in a distinct and transient increase of the cytosolic Ca2+ concentration. This Ca2+ peak was reduced by the T-type channel blocker, Ni2+, by the L-type channel blockers, nifedipine and verapamil, and by the N-type channel blocker, ,-conotoxin GVIA. The P/Q-type channel antagonist, ,-agatoxin TK, had no effect on postsynaptic [Ca2+]i. The NMDA receptor channel blocker D-AP5 reduced the Ca2+ peak, whereas the AMPA receptor channel blocker CNQX had no effect. Postsynaptic currents, monosynaptically evoked by electrical stimulation of the attached dorsal roots with C-fibre and A,-fibre intensity, respectively, were reduced by N-type channel blocker ,-conotoxin GVIA and to a much lesser extent, by P/Q-type channel antagonist ,-agatoxin TK, and the L-type channel blockers verapamil, respectively. No difference was found between unidentified neurons and neurons projecting to the periaqueductal grey matter. This is the first quantitative description of the relative contribution of voltage-dependent Ca2+ channels to the synaptic transmission in lamina I of the spinal dorsal horn, which is essential in the processing of pain-related information in the central nervous system. [source] Trimetazidine Reduces Basal Cytosolic Ca2+ Concentration During Hypoxia in Single Xenopus Skeletal MyocytesEXPERIMENTAL PHYSIOLOGY, Issue 3 2003C. M. Stary We tested the hypotheses that: (1) Ca2+ handling and force production would be irreversibly altered in skeletal muscle during steady-state contractions when subjected to severe, prolonged hypoxia and subsequent reoxygenation; and (2) application of the cardio-protective drug trimetazidine would attenuate these alterations. Single, living skeletal muscle fibres from Xenopus laevis were injected with the Ca2+ indicator fura 2, and incubated for 1 h prior to stimulation in 100 ,M TMZ-Ringer solution (TMZ; n = 6) or standard Ringer solution (CON; n = 6). Force and relative free cytosolic Ca2+ concentration ([Ca2+]c) were measured during continuous tetanic contractions produced every 5 s as fibres were sequentially perfused in the following manner: 3 min high extracellular PO2 (159 mmHg), 15 min hypoxic perfusion (3-5 mmHg) then 3 min high PO2. Hypoxia caused a decrease in force and peak [Ca2+]c in both the TMZ and CON fibres, with no significant (P < 0.05) difference between groups. However, basal [Ca2+]c was significantly lower during hypoxia in the TMZ group vs. the CON group. While reoxygenation generated only modest recovery of relative force and peak [Ca2+]c in both groups, basal [Ca2+]c remained significantly less in the TMZ group. These results demonstrated that in contracting, single skeletal muscle fibres, TMZ prevented increases in basal [Ca2+]c generated during a severe hypoxic insult and subsequent reoxygenation, yet failed to protect the cell from the deleterious effects of prolonged hypoxia followed by reoxygenation. [source] The effect of short-term low-temperature treatments on gene expression in Arabidopsis correlates with changes in intracellular Ca2+ levelsPLANT CELL & ENVIRONMENT, Issue 4 2003K. NORDIN HENRIKSSON ABSTRACT The role of changes in intracellular calcium ion concentration ([Ca2+]i) in low-temperature signal transduction in plants has lately been supported by several studies. An analysis to determine whether the low-temperature-induced increase in cytosolic Ca2+ concentration ([Ca2+]cyt) could be correlated with a downstream response such as gene expression was carried out. The induction of the low-temperature-regulated gene LTI78 was used as an end point marker of the signal transduction pathway. It was found that this gene is induced by very brief low-temperature exposures and that the induction does not depend on a continuous exposure to low temperature. By altering the cooling rate, different patterns of the Ca2+ response were obtained which could be correlated with different patterns of LTI78 induction. Furthermore, reducing the Ca2+ transients by pre-treatment with the Ca2+ channel blocker La3+ also led to a reduced level of gene induction. The results show that brief exposures to low temperature results in the onset of a signalling pathway that leads to the induction of gene expression. This indicates the involvement of changes in [Ca2+]cyt in low-temperature signalling leading to LTI78 expression but the presence of multiple signalling pathways is suggested. [source] Cytosolic Ca2+ concentration and rate of increase of the cytosolic Ca2+ concentration in the regulation of vascular permeability in Rana in vivoTHE JOURNAL OF PHYSIOLOGY, Issue 3 2005C. A. Glass Vascular permeability is assumed to be regulated by the cytosolic Ca2+ concentration ([Ca2+]c) of the endothelial cells. When permeability is increased, however, the maximum [Ca2+]c appears to occur after the maximum permeability increase, suggesting that Ca2+ -dependent mechanisms other than the absolute Ca2+ concentration may regulate permeability. Here we investigate whether the rate of increase of the [Ca2+]c (d[Ca2+]c/dt) may more closely approximate the time course of the permeability increase. Hydraulic conductivity (Lp) and endothelial [Ca2+]c were measured in single perfused frog mesenteric microvessels in vivo. The relationships between the time courses of the increased Lp, [Ca2+]c and d[Ca2+]c/dt were examined. Lp peaked significantly earlier than [Ca2+]c in all drug treatments examined (Ca2+ store release, store-mediated Ca2+ influx, and store-independent Ca2+ influx). When Lp was increased in a store-dependent manner the time taken for Lp to peak (3.6 ± 0.9 min during store release, 1.2 ± 0.3 min during store-mediated Ca2+ influx) was significantly less than the time taken for [Ca2+]c to peak (9.2 ± 2.8 min during store release, 2.1 ± 0.7 min during store-mediated influx), but very similar to that for the peak d[Ca2+]c/dt to occur (4.3 ± 2.0 min during store release, 1.1 ± 0.5 min during Ca2+ influx). Additionally, when the increase was independent of intracellular Ca2+ stores, Lp (0.38 ± 0.03 min) and d[Ca2+]c/dt (0.30 ± 0.1 min) both peaked significantly before the [Ca2+]c (1.05 ± 0.31 min). These data suggest that the regulation of vascular permeability by endothelial cell Ca2+ may be regulated by the rate of change of the [Ca2+]c rather than the global [Ca2+]. [source] Receptor signaling mechanisms underlying muscarinic agonist-evoked contraction in guinea-pig ileal longitudinal smooth muscleBRITISH JOURNAL OF PHARMACOLOGY, Issue 2 2003T Unno In guinea-pig ileal longitudinal muscle, muscarinic partial agonists, 4-(N -[3-chlorophenyl]-carbomoyloxy)-2-butynyl-trimethylammonium (McN-A343) and pilocarpine, each produced parallel increases in tension and cytosolic Ca2+ concentration ([Ca2+]c) with a higher EC50 than that of the full agonist carbachol. The maximum response of [Ca2+]c or tension was not much different among the three agonists. The Ca2+ channel blocker nicardipine markedly inhibited the effects of all three agonists The contractile response to any agonist was antagonized in a competitive manner by M2 receptor selective antagonists (N,N,- bis[6-[[(2-methoyphenyl)methyl]amino]hexyl]-1,8-octanediamine tetrahydrochloride and 11-[[2-[(diethlamino)methyl]-1-piperidinyl]acetyl]-5,11-dihydro-6H-pyrido[2,3-b][1,4] benzodiazepine-6-one), and the apparent order of M2 antagonist sensitivity was McN-A343>pilocarpine>carbachol. M3 receptor selective antagonists, 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide and darifenacin, both severely depressed the maximum response for McN-A343, while darifenacin had a similar action in the case of pilocarpine. Both M3 antagonists behaved in a competitive manner in the case of the carbachol response. McN-A343 failed to release Ca2+ from the intracellular stores, and the Ca2+ -releasing action of pilocarpine was very weak compared with that of carbachol. All three agonists were capable of increasing Ca2+ sensitivity of the contractile proteins. McN-A343 rarely produced membrane depolarization, but always accelerated electrical spike discharge. Pilocarpine effect was more often accompanied by membrane depolarization, as was usually seen using carbachol. The results suggest that muscarinic agonist-evoked contractions result primarily from the integration of Ca2+ entry associated with the increased spike discharge and myofilaments Ca2+ sensitization, and that Ca2+ store release may contribute to the contraction indirectly via potentiation of the electrical membrane responses. They may also support the idea that an interaction of M2 and M3 receptors plays a crucial role in mediating the contraction response. British Journal of Pharmacology (2003) 139, 337,350. doi:10.1038/sj.bjp.0705267 [source] Walker tumor cells express larger amounts of the antiapoptotic protein Bcl-2 and presents higher resistance to toxic concentrations of Ca2+ than the tumor cells K 562DRUG DEVELOPMENT RESEARCH, Issue 4 2001Graziela Milani Abstract Ca2+ homeostasis was studied in two tumor cell lines (Walker 256 and K 562) previously shown to exhibit different mitochondrial Ca2+ accumulation capacity. When intact, both cells present cytosolic Ca2+ concentrations within the range expected for mammalian cells, as determined through fura-2 fluorescence ratios. In order to study intracellular Ca2+ distribution, digitonin was used to permeabilize the plasma membrane without affecting intracellular organelle structure, as assessed using electron microscopy. Digitonin-permeabilized Walker 256 cells incubated with Ca2+ presented uptake of the cation exclusively through mitochondrial activity. In addition, very large Ca2+ loads were necessary to promote a disruption of Walker 256 mitochondrial membrane potential. K 562 cells presented active Ca2+ uptake through both nonmitochondrial and mitochondrial compartments and suffered disruption of mitochondrial membrane potential at lower Ca2+ loads than Walker 256 mitochondria. The higher Ca2+ resistance in Walker 256 cells could be attributed to Bcl-2 overexpression, as evidenced by immunocytochemical staining. Thus, we correlate natural Bcl-2 overexpression, observed in Walker 256 cells, with higher resistance to mitochondrial Ca2+ overload, as was shown previously in mitochondria from cells transfected with the bcl-2 gene. Drug Dev. Res. 52:508,514, 2001. © 2001 Wiley-Liss, Inc. [source] ,-Latrotoxin increases spontaneous and depolarization-evoked exocytosis from pancreatic islet ,-cellsTHE JOURNAL OF PHYSIOLOGY, Issue 3 2005Amelia M. Silva ,-Latrotoxin (,-LT), a potent excitatory neurotoxin, increases spontaneous, as well as action potential-evoked, quantal release at nerve terminals and increases hormone release from excitable endocrine cells. We have investigated the effects of ,-LT on single human, mouse and canine ,-cells. In isolated and combined measurements, ,-LT, at nanomolar concentrations, induces: (i) rises in cytosolic Ca2+, into the micromolar range, that are dependent on extracellular Ca2+; (ii) large conductance non-selective cation channels; and (iii) Ca2+ -dependent insulin granule exocytosis, measured as increases in membrane capacitance and quantal release of preloaded serotonin. Furthermore, at picomolar concentrations, ,-LT potentiates depolarization-induced exocytosis often without evidence of inducing channel activity or increasing cytosolic Ca2+. These results strongly support the hypothesis that ,-LT, after binding to specific receptors, has at least two complementary modes of action on excitable cells. (i) ,-LT inserts into the plasma membrane to form Ca2+ permeable channels and promote Ca2+ entry thereby triggering Ca2+ -dependent exocytosis in unstimulated cells. (ii) At lower concentrations, where its channel forming activity is hardly evident, ,-LT augments depolarization-evoked exocytosis probably by second messenger-induced enhancement of the efficiency of the vesicle recruitment or vesicle fusion machinery. We suggest that both modes of action enhance exocytosis from a newly described highly Ca2+ -sensitive pool of insulin granules activated by global cytosolic Ca2+ concentrations in the range of ,1 ,m. [source] Itraconazole-mediated inhibition of calcium entry into platelet-activating factor-stimulated human neutrophils is due to interference with production of leukotriene B4CLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 1 2007H. C. Steel Summary The primary objective of this study was to probe the involvement of leukotriene B4 (LTB4) in itraconazole (0·1,5 µM)-mediated inhibition of Ca2+ uptake by chemoattractant-activated human neutrophils. Following exposure of the cells to platelet-activating factor (PAF, 200 nM), LTB4 was measured by immunoassay, while neutrophil cytosolic Ca2+ concentrations were determined by a fura-2/AM-based spectrofluorimetric procedure. Activation of neutrophils was accompanied by an abrupt and sustained (for about 1 min) elevation in cytosolic Ca2+ which was associated with increased generation of LTB4, both of which were attenuated significantly by itraconazole at 0·5 µM and higher. The inhibitory effect of the anti-mycotic on Ca2+ uptake by PAF-activated cells was mimicked by an LTB4 antibody, as well as by LY255283 (1 µM) and MK886 (0·5 µM), an antagonist of LTB4 receptors and an inhibitor of 5,-lipoxygenase-activating protein, respectively, while addition of itraconazole to purified 5,-lipoxygenase resulted in inhibition of enzyme activity. A mechanistic relationship between itraconazole-mediated inhibition of LTB4 production and Ca2+ influx was also supported by the observation that pulsed addition of purified LTB4 to PAF-activated neutrophils caused substantial restoration of Ca2+ uptake by cells treated with the anti-mycotic. Taken together, these observations suggest that the potentially beneficial anti-inflammatory interactions of itraconazole with activated neutrophils result from interference with production of LTB4, with consequent attenuation of a secondary LTB4 -mediated wave of Ca2+ uptake by the cells. [source] | |||||||||||||