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1.
Gerasimenko O  Tepikin A 《Cell calcium》2005,38(3-4):201-211
The review will aim to briefly summarise information on calcium measurements in cellular organelles with emphases on studies conducted in live cells using optical probes. When appropriate we will try to compare the effectiveness of different indicators for intraorganellar calcium measurements. We will consider calcium measurements in endoplasmic reticulum, Golgi apparatus, endosomes/lysosomes, nucleoplasm, nuclear envelope, mitochondria and secretory granules.  相似文献   

2.
Pigment aggregation in melanophores of Labrus ossifagus is controlled by an alpha2-adrenoceptor and is somehow modulated by melatonin. The signal transduction mechanisms seem to involve both an attenuation of cAMP and an increase in intracellular Ca2+, inhibiting protein kinase A or activating a phosphatase, respectively. These effects result in dephosphorylation, which in turn induces aggregation. Various alpha2-adrenoceptor agonists attenuate cAMP levels or increase the concentration of intracellular Ca2+. Noradrenaline, for example, lowers cAMP but does not affect the calcium signal whereas B-HT 920, an alpha2-adrenoceptor specific agonist, does not induce a cAMP decrease but does appear to induce an increase in intracellular Ca2+. This later inference is drawn from experiments with BAPTA/AM, an intracellular calcium chelator, which counteracts the aggregation induced by B-HT 920. Interestingly, the very potent alpha2-adrenoceptor agonist medetomidine apparently activates both signal transduction pathways, which could explain its high efficacy in producing aggregation. Melatonin itself does not cause pigment aggregation, but it potentiates noradrenaline-induced aggregation. It has been suggested that melatonin receptors and alpha2-adrenoceptors follow the same signal transduction pathway, i.e. an attenuation of cAMP. In our experiments, melatonin did not reduce cAMP levels; instead it appears to increase Ca2+ concentration, since melatonin-potentiated aggregation was inhibited by BAPTA/AM. Thus, aggregation amplified by melatonin is probably not mediated by a further decrease in cAMP, but by the same signal transduction mechanism as B-HT 920, i.e. an increase in Ca2+. This further strengthens the suggestion that melatonin and B-HT 920 bind to the same site, but it is unclear if that particular site is on the melatonin receptor or the alpha2-adrenoceptor.  相似文献   

3.
《Life sciences》1993,53(8):PL135-PL140
Activation of the α1-adrenoceptor (α1-AR) in mouse peritoneal macrophages induces a Ca2+-dependent K+ current via release of Ca2+ from internal stores [Hara et al. (1991) Pflügers Arch 419: 371–379]. In order to characterize the α1-AR subtype in macrophages, we examined potencies of the α1-AR subtype-selective antagonists against epinephrine-induced increases in intracellular Ca2+ concentration ([Ca2+]i). The Ki values for the α1-AR in macrophages of WB4101, 5-methylurapidil and phentolamine were 0.13 nM, 0.15 nM and 7.0 nM, respectively. (+)-Niguldipine (10 nM) completely blocked the response. Treatment with chloroethylclonidine (CEC) did not abolish the response. These pharmacological properties of the α1-AR in macrophages agree quite well with those of the α1A subtype.  相似文献   

4.
Effect of endothelin-1 and chemically induced hypoxia on Na+−K+−Cl cotransport activity in cultured rat brain capillary endothelial cells was examined by using86Rb+ as a tracer for K+; bumetanide-sensitive K+ uptake was defined as Na+−K+−Cl cotransport activity. Endothelin-1, phorbol 12-myristate 13-acetate (PMA), or thapsigargin increased Na+−K+−Cl cotransport activity. A protein kinase C inhibitor, bisindolylmaleimide, inhibited PMA- and endothelin-1- (but not thapsigargin-) induced Na+−K+−Cl cotransport activity, indicating the presence of both protein kinase C-dependent regulatory mechanisms and protein kinase C-independent mechanisms which involve intracellular Ca2+. Oligomycin, sodium azide, or antimycin A increased Na+−K+−Cl cotransport activity by 80–200%. Oligomycin-induced Na+−K+−Cl cotransport activity was reduced by an intracellular Ca2+ chelator (BAPTA/AM) but not affected by bisindolylmaleimide, suggesting the involvement of intracellular Ca2+, and not protein kinase C, in hypoxia-induced Na+−K+−Cl cotransport activity. Portions were presented at “27th Annual Meeting, The American Society for Neurochemistry” Philadelphia, Pennsylvania, March 2–6, 1996.  相似文献   

5.
By the same method as in the previous paper by the author (Bull. Math. Biophysics,23, 55–68, 1961), the mobile electron system is studied in tryptophan which has donated an electron to another molecule. The obtained net charge distribution of tryptophan+ is compared with those of the semiquinones of riboflavin (FMN, FMNH and FMNH 2 + ). From this comparison, a suggestion is obtained for the structure of each of two charge transfer complexes between riboflavin and tryptophan in neutral and acid solution.  相似文献   

6.
α-Lactalbumin possesses multiple Zn2+ binding sites, with the strongest site having an affinity constant of 5×105 M?1 [Permyakovet al. (1991),J. Protein Chem. 100, 577]. The binding of zinc at secondary sites is accompanied by destabilization of the protein structure and progressive protein aggregation. This pronounced destabilization is reflected in a shift of the thermal denaturation transition temperature by more than 40°. The present work examines Co2+ binding to bovineα-lactalbumin, where for this analog of Zn2+, multiple binding sites were also found from spectrofluorimetric titrations. The strong site Co2+ binding constant was 1.3×106 M?1. However, in contrast to Zn2+ binding, Co2+ does not cause protein aggregation nor any significant thermal destabilization of the protein. Fluroescence energy transfer measurements between Tb3+ in the strong calcium site to Co2+ in the strong Zn2+ site gave a distance in the range of 14–18 Å, which was in excellent agreement with recent crystallographic data for humanα-lactalbumin [Renet al. (1993), J. Biol. Chem.268, 19292–19298] However, the X-ray structure did not identify the additional zinc sites found from earlier solution studies, presumably due to restrictive crystal packing interactions. The results from the current work confirm that the strong cobalt (zinc) site in solution is the same zinc site elucidated by X-ray crystallography.  相似文献   

7.
Chen M  Fernandez HL 《Cell calcium》1999,26(3-4):149-154
Ca2+ signaling is a central process in brain function, but the direction of its change in ageing and in the presymptomatic stages of Alzheimer's disease has been controversial. A great deal of studies have been interpreted as supportive to the current hypothesis that intracellular Ca2+ levels are steadily increased in ageing. We, however, believe that, although current studies have provided valuable knowledge for the mechanisms of the signal transduction process, they have not furnished relevant information regarding the global Ca2+ changes in the ageing brain, because the cognition-related Ca2+ pulses exist only in the intact brain.  相似文献   

8.
Macrophage polarization to an anti-inflammatory phenotype upon contact with apoptotic cells is a contributing hallmark to immune suppression during the late phase of sepsis. Although the peroxisome proliferator-activated receptor γ (PPARγ) supports this macrophage phenotype switch, it remains elusive how apoptotic cells activate PPARγ. Assuming that a molecule causing PPARγ activation in macrophages originates in the cell membrane of apoptotic cells we analyzed lipid rafts from apoptotic, necrotic, and living human Jurkat T cells which showed the presence of 5-lipoxygenase (5-LO) in lipid rafts of apoptotic cells only. Incubating macrophages with lipid rafts of apoptotic, but not necrotic or living cells, induced PPAR responsive element (PPRE)-driven mRuby reporter gene expression in RAW 264.7 macrophages stably transduced with a 4xPPRE containing vector. Experiments with lipid rafts of apoptotic murine EL4 T cells revealed similar results. To verify the involvement of 5-LO in activating PPARγ in macrophages, Jurkat T cells were incubated with the 5-LO inhibitor MK-866 prior to induction of apoptosis, which failed to induce mRuby expression. Similar results were obtained with lipid rafts of apoptotic EL4 T cells preexposed to the 5-LO inhibitors zileuton and CJ-13610. Interestingly, Jurkat T cells overexpressing 5-LO failed to activate PPARγ in macrophages, while their 5-LO overexpressing apoptotic counterparts did. Our results suggest that during apoptosis 5-LO gets associated with lipid rafts and synthesizes ligands that in turn stimulate PPARγ in macrophages.  相似文献   

9.
Increasing evidence suggests that the ubiquitin-binding histone deacetylase-6 (HDAC6) plays an important role in the clearance of misfolded proteins by autophagy. In this study, we treated PC-12 cells over-expressing human mutant (A53T) α-synuclein (α-syn) and SH-SY5Y cells with MPP(+). It was found that HDAC6 expression significantly increased and mainly colocalized with α-syn in the perinuclear region to form aggresome-like bodies. HDAC6 deficiency blocked the formation of aggresome-like bodies and interfered with the autophagy in response to MPP(+)-induced stress. Moreover, misfolded α-syn accumulated into the nuclei, resulting in its reduced clearance, and finally, the number of apoptotic cells significantly increased. Taken together, HDAC6 participated in the degradation of MPP(+)-induced misfolded α-syn aggregates by regulating the aggresome-autophagy pathway. Understanding the mechanism may disclose potential therapeutic targets for synucleinopathies such as Parkinson's disease.  相似文献   

10.
Apoptosis - Melanoma is an aggressive tumor with still poor therapy outcomes. δ-tocotrienol (δ-TT) is a vitamin E derivative displaying potent anti-cancer properties. Previously, we...  相似文献   

11.
We previously showed that injury by partial duct ligation (PDL) in adult mouse pancreas activates Neurogenin 3 (Ngn3)+ progenitor cells that can differentiate to β cells ex vivo. Here we evaluate the role of Ngn3+ cells in β cell expansion in situ. PDL not only induced doubling of the β cell volume but also increased the total number of islets. β cells proliferated without extended delay (the so-called ‘refractory'' period), their proliferation potential was highest in small islets, and 86% of the β cell expansion was attributable to proliferation of pre-existing β cells. At sufficiently high Ngn3 expression level, upto 14% of all β cells and 40% of small islet β cells derived from non-β cells. Moreover, β cell proliferation was blunted by a selective ablation of Ngn3+ cells but not by conditional knockout of Ngn3 in pre-existing β cells supporting a key role for Ngn3+ insulin cells in β cell proliferation and expansion. We conclude that Ngn3+ cell-dependent proliferation of pre-existing and newly-formed β cells as well as reprogramming of non-β cells contribute to in vivo β cell expansion in the injured pancreas of adult mice.  相似文献   

12.
The chronic effects of type 2 diabetes mellitus on myofilament sensitivity to Ca(2+) in ventricular myocytes from the Goto-Kakizaki (GK) rat have been investigated. Experiments were performed in ventricular myocytes isolated from 17-month GK rats and age-matched Wistar controls. Myocytes were loaded with fura-2 (an indicator for intracellular Ca(2+) concentration) and the fura-2 ratio (340/380 nm), and shortening were measured simultaneously in electrically stimulated myocytes. Myofilament sensitivity to Ca(2+) was assessed from phase-plane diagrams of fura-2 versus cell length by measuring the gradient of the fura-2-cell length trajectory during late relaxation of the twitch contraction. Non-fasting and fasting blood glucose were elevated in GK rats compared to controls. Fasting blood glucose was 151.5 +/- 15.3 mg/dl (n = 8) in GK rats compared to 72.1 +/- 3.6 mg/dl (n = 9) in controls. At 120 min after intraperitoneal injection of glucose (2 g/kg body weight), blood glucose was 570.8 +/- 36.8 mg/dl in GK rats compared to 148 +/- 8.6 mg/dl in controls. Amplitude of shortening was significantly increased in myocytes from GK rats (6.56 +/- 0.54%, n = 31) compared to controls (5.05 +/- 0.43%, n = 36), and the amplitude of the Ca(2+) transient was decreased in myocytes from GK rats (0.23 +/- 0.02 RU, n = 31) compared to controls (0.30 +/- 0.02 RU, n = 36). The fura-2-cell length trajectory during the late stages of relaxation of the twitch contraction was steeper in myocytes from GK rats (89.2 +/- 16.6 microm/RU, n = 27) compared to controls (31.9 +/- 5.9 microm/RU, n = 35). Increased amplitude of shortening, accompanied by a decrease in amplitude of the Ca(2+) transient, might be explained by an increased myofilament sensitivity to Ca(2+).  相似文献   

13.
Besides CD4+CD25+Foxp3+ regulatory T cells (Tregs), other immunosuppressive T cells also participated in the regulation of immune tolerance. Reportedly, neuropilin-1 (Nrp1) might be one of the molecules by which regulatory cells exert their suppressive effects. Indeed, CD4+CD25Nrp1+ T cells exhibit potent suppressive function in autoimmune inflammatory responses. Here we investigated the specific role of CD4+CD25Nrp1+ T cells in the setting of the transplant immune response. Through MLR assays, we found that CD4+CD25Nrp1+ T cells suppressed the proliferation of naive CD4+CD25 T cells activated by allogeneic antigen-stimulation. Adoptive transfer of CD4+CD25Nrp1+ T cells synergized with rapamycin to induce long-term graft survival in fully MHC-mismatched murine heart transplantation, which was associated with decreased IFN-γ, IL-17 and increased IL-10, TGF-β, Foxp3 and Nrp1 expression in the grafts. Importantly, our data indicated that CD4+CD25Nrp1+ T cell transfer augments the accumulation of Tregs in the recipient, and creates conditions that favored induction of hyporesponsiveness of the T effector cells. In conclusion, this translational study indicates the possible therapeutic potential of CD4+CD25Nrp1+ T cells in preventing allorejection. CD4+Nrp1+ T cells might therefore be used in bulk as a population of immunosuppressive cells with more beneficial properties concerning ex vivo isolation as compared to Foxp3+ Tregs.  相似文献   

14.
15.
We demonstrate that the application of the muscarinic agonist carbachol to the human salivary epithelial cell line HSG elicits the now well-known phenomenon of quantal Ca2+-release; namely, that the application of a submaximal concentration of agonist results in the release of only a portion of the agonist-sensitive intracellular Ca2+ pool. One explanation that has been proposed to account for this effect is that there are multiple intracellular Ca2+ stores, each with a different agonist sensitivity, which release Ca2+ in an all-or-none fashion. We test this hypothesis in intact HSG cells with an experimental protocol designed to preferentially load less-agonist-sensitive stores with 45Ca2+ and more-agonist-sensitive stores with 45Ca2+. However, contrary to the expectations of the above explanation, these cells do not preferentially release 45Ca2+ in response to low concentrations of agonist. Thus our data suggest that quantal Ca2+ release must arise from some other property of the stores or their Ca2+ release channels.  相似文献   

16.
17.
Although synergies between reducing emissions from deforestation and forest degradation in developing countries (REDD+) and biodiversity conservation are generally expected, they will not be achieved through poorly designed REDD+ schemes. Perceptions of actors in REDD+ implementation processes can guide the design of appropriate policies. Semi-structured interviews were conducted in Peru with actors in REDD+ working groups at the national and subnational level (n = 50) to explore how they relate REDD+ implementation to biodiversity conservation and other policy issues, and to identify the aspects of biodiversity they regard as especially important (biodiversity conservation values). Interviews were analysed using qualitative content analysis; actors’ statements were summarised into nine policy issue categories. Issues raised often were related to governance, socioeconomic aspects and land management in REDD+ implementation (47 % of the statements). Biodiversity issues were mentioned less often (23 %). Direct synergies between REDD+ and biodiversity conservation were assumed by most actors and few concrete risks and corresponding safeguards were mentioned. Actors mentioned 177 biodiversity conservation values, which were classified into different value types. Most values were aspects of biodiversity with direct or indirect use values (68 %). Aspects which potentially provide future benefits (option values) or resilience values were mentioned less often (5 and 16 %, respectively). The opportunity to foster conservation of use values as additional benefits from REDD+ is generally recognised by the actors, but aspects of biodiversity that are important for the long-term integrity of forest ecosystems received less attention. It is thus important to push for the integration of respective safeguards in REDD+ strategies.  相似文献   

18.
The epithelial Na(+) channel (ENaC) is a heteromultimeric ion channel that plays a key role in Na(+) reabsorption across tight epithelia. The canonical ENaC is formed by three analogous subunits, α, β, and γ. A fourth ENaC subunit, named δ, is expressed in the nervous system of primates, where its role is unknown. The human δ-ENaC gene generates at least two splice isoforms, δ(1) and δ(2) , differing in the N-terminal sequence. Neurons in diverse areas of the human and monkey brain differentially express either δ(1) or δ(2) , with few cells coexpressing both isoforms, which suggests that they may play specific physiological roles. Here we show that heterologous expression of δ(1) in Xenopus oocytes and HEK293 cells produces higher current levels than δ(2) . Patch-clamp experiments showed no differences in single channel current magnitude and open probability between isoforms. Steady-state plasma membrane abundance accounts for the dissimilarity in macroscopic current levels. Differential trafficking between isoforms is independent of β- and γ-subunits, PY-motif-mediated endocytosis, or the presence of additional lysine residues in δ(2)-N terminus. Analysis of δ(2)-N terminus identified two sequences that independently reduce channel abundance in the plasma membrane. The δ(1) higher abundance is consistent with an increased insertion rate into the membrane, since endocytosis rates of both isoforms are indistinguishable. Finally, we conclude that δ-ENaC undergoes dynamin-independent endocytosis as opposed to αβγ-channels.  相似文献   

19.
The mitogen-activated protein kinase (MAPK) pathway is a highly conserved signaling cascade that converts extracellular signals into various outputs. In Caenorhabditis elegans, asymmetric expression of the candidate odorant receptor STR-2 in either the left or the right of two bilaterally symmetrical olfactory AWC neurons is regulated by axon contact and Ca2+ signaling. We show that the MAPK kinase (MAPKK) SEK-1 is required for asymmetric expression in AWC neurons. Genetic and biochemical analyses reveal that SEK-1 functions in a pathway downstream of UNC-43 and NSY-1, Ca2+/calmodulin-dependent protein kinase II (CaMKII) and MAPK kinase kinase (MAPKKK), respectively. Thus, the NSY-1–SEK-1–MAPK cascade is activated by Ca2+ signaling through CaMKII and establishes asymmetric cell fate decision during neuronal development.  相似文献   

20.
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