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Perez-Alvarez S.,University of Castilla - La Mancha | Solesio M.E.,Albacete University Hospital Center | Cuenca-Lopez M.D.,University of Castilla - La Mancha | Melero-Fernndez De Mera R.M.,University of Castilla - La Mancha | And 5 more authors.
International Journal of Cell Biology | Year: 2012

Previously, we have shown that SH-SY5Y cells exposed to high concentrations of methadone died due to a necrotic-like cell death mechanism related to delayed calcium deregulation (DCD). In this study, we show that, in terms of their Ca2+ responses to 0.5mM methadone, SH-SY5Y cells can be pooled into four different groups. In a broad pharmacological survey, the relevance of different Ca2+-related mechanisms on methadone-induced DCD was investigated including extracellular calcium, L-type Ca2+ channels, -opioid receptor, mitochondrial inner membrane potential, mitochondrial ATP synthesis, mitochondrial Ca2+/2Na+-exchanger, reactive oxygen species, and mitochondrial permeability transition. Only those compounds targeting mitochondria such as oligomycin, FCCP, CGP 37157, and cyclosporine A were able to amend methadone-induced Ca2+ dyshomeostasis suggesting that methadone induces DCD by modulating the ability of mitochondria to handle Ca2+. Consistently, mitochondria became dramatically shorter and rounder in the presence of methadone. Furthermore, analysis of oxygen uptake by isolated rat liver mitochondria suggested that methadone affected mitochondrial Ca2+ uptake in a respiratory substrate-dependent way. We conclude that methadone causes failure of intracellular Ca2+ homeostasis, and this effect is associated with morphological and functional changes of mitochondria. Likely, this mechanism contributes to degenerative side effects associated with methadone treatment. Copyright © 2012 Sergio Perez-Alvarez et al. Source

Montero O.,Center for Biotechnology Development | Sanchez-Guijo A.,Institute of Molecular Biology and Genetics IBGM | Lubian L.M.,CSIC - Institute of Marine Sciences | Martinez-Rodriguez G.,CSIC - Institute of Marine Sciences
Journal of Biosciences | Year: 2012

Time course of carotenoid and membrane lipid variation during high light (HL) acclimation (about 85 μmol m-2 s-1), after transfer from low light (LL) (5-10 μmol m-2 s-1), was determined in a marine Synechococcus strain. High-performance liquid chromatography (HPLC) coupled to diode array detector (DAD) or electrospray ionization mass spectrometry (ESI-MS) was used for compound separation and detection. Myxoxanthophyll rose within a time interval of 8 h to 24 h after the onset of exposure to HL. β-carotene content started to decrease after 4 h of the onset of exposure to HL. Zeaxanthin content rose with exposure to HL, but it was only significant after 24 h of exposure. Carotenoid changes are in agreement with a coordinated activity of the enzymes of the myxoxanthophyll biosynthetic pathway, with no rate-limiting intermediate steps. Lipid analysis showed all species with a C18:3/C16:0 composition increased their content, the changes of PG(18:3/16:0) and MGDG(18:3/16:0) being primarily significant. Major lipid changes were also found to occur within 24 h. These changes might suggest reduction and reorganization of the thylakoid membrane structure. Hypotheses are also drawn on the role played by lipid molecule shape and their possible effect in membrane fluidity and protein accommodation. © Indian Academy of Sciences. Source

Sobradillo D.,Institute of Molecular Biology and Genetics IBGM | Hernandez-Morales M.,Institute of Molecular Biology and Genetics IBGM | Ubierna D.,Institute of Molecular Biology and Genetics IBGM | Moyer M.P.,INCELL Corporation | And 2 more authors.
Journal of Biological Chemistry | Year: 2014

Results: Colon carcinoma cells show enhanced store-operated Ca2+ entry and currents and depleted Ca2+ stores associated with changes in STIM1/STIM2 ratio and TRPC1.Conclusion: Ca2+ remodeling in colon cancer is driven by a reciprocal shift in TRPC1 and STIM2.Significance: STIM1/STIM2 and TRPC1 should be investigated further as novel targets for colon cancer.Background: Changes in Ca2+handling in tumor cells might provide novel targets for cancer. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc. Source

Munoz E.,Institute of Molecular Biology and Genetics IBGM | Hernandez-Morales M.,Institute of Molecular Biology and Genetics IBGM | Sobradillo D.,Institute of Molecular Biology and Genetics IBGM | Rocher A.,University of Valladolid | And 2 more authors.
Cell Calcium | Year: 2013

Vascular smooth muscle cells undergo phenotypic switches after damage which may contribute to proliferative disorders of the vessel wall. This process has been related to remodeling of Ca2+ channels. We have tested the ability of cultured human coronary artery smooth muscle cells (hCASMCs) to return from a proliferative to a quiescent behavior and the contribution of intracellular Ca2+ remodeling to the process. We found that cultured, early passage hCASMCs showed a high proliferation rate, sustained increases in cytosolic [Ca2+] in response to angiotensin II, residual voltage-operated Ca2+ entry, increased Stim1 and enhanced store-operated currents. Non-steroidal anti-inflammatory drugs inhibited store-operated Ca2+ entry and abolished cell proliferation in a mitochondria-dependent manner. After a few passages, hCASMCs turned to a quiescent phenotype characterized by lack of proliferation, oscillatory Ca2+ response to angiotensin II, increased Ca2+ store content, enhanced voltage-operated Ca2+ entry and Cav1.2 expression, and decreases in Stim1, store-operated current and store-operated Ca2+ entry. We conclude that proliferating hCASMCs return to quiescence and this switch is associated to a remodeling of Ca2+ channels and their control by subcellular organelles, thus providing a window of opportunity for targeting phenotype-specific Ca2+ channels involved in proliferation. © 2013 Elsevier Ltd. Source

Caballero E.,Institute of Molecular Biology and Genetics IBGM | Calvo-Rodriguez M.,Institute of Molecular Biology and Genetics IBGM | Gonzalo-Ruiz A.,University of Valladolid | Villalobos C.,Institute of Molecular Biology and Genetics IBGM | And 2 more authors.
Neuroscience Letters | Year: 2016

Oligomers of the amyloid β peptide (Aβo) are becoming the most likely neurotoxin in Alzheimer's disease. Controversy remains on the mechanisms involved in neurotoxicity induced by Aβo and the targets involved. We have reported that Aβo promote Ca2+ entry, mitochondrial Ca2+ overload and apoptosis in cultured cerebellar neurons. However, recent evidence suggests that some of these effects could be induced by glutamate receptor agonists solved in F12, the media in which Aβo are prepared. Here we have tested the effects of different media on Aβo formation and on cytosolic Ca2+ concentration ([Ca2+]cyt) in rat cerebellar and hippocampal cell cultures. We found that Aβo prepared according to previous protocols but solved in alternative media including saline, MEM and DMEM do not allow oligomer formation and fail to increase [Ca2+]cyt. Changes in the oligomerization protocol and supplementation of media with selected salts reported to favor oligomer formation enable Aβo formation. Aβo prepared by the new procedure and containing small molecular weight oligomers increased [Ca2+]cyt, promoted mitochondrial Ca2+ overload and cell death in cerebellar granule cells and hippocampal neurons. These results foster a role for Ca2+ entry in neurotoxicity induced by Aβo and provide a reliable procedure for investigating the Ca2+ entry pathway promoted by Aβo. © 2015 Elsevier Ireland Ltd. Source

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