1,720,998 research outputs found
Fluoxetine prevents acetylcholine-induced excitotoxicity blocking human endplate acetylcholine receptor.
No full textIntroduction: Fluoxetine is an open channel blocker of fetal muscle acetylcholine (ACh) receptor (AChR) and slow-channel mutant AChRs. It is used commonly to treat patients with slow-channel congenital myasthenic syndromes. Fluoxetine effects on adult wild-type endplate AChR are less characterized, although muscle AChR isoforms are differentially modulated by some drugs. Methods: Excitotoxicity assays and patch clamp recordings were performed in human embryonic kidney 293 (HEK) cells expressing wild-type or slow-channel mutant human AChRs. Results: Fluoxetine (2-10 μM) abolished ACh-induced death and decreased ACh-activated whole-cell currents in cells expressing all AChR types. In outside-out patches, fluoxetine rapidly curtailed ACh evoked unitary activity and macroscopic currents. The effect was increased if fluoxetine was applied before ACh. Conclusions: Fluoxetine is an open channel blocker, but it also affects AChR in the closed state. AChR blockade likely underlies the rescue of HEK cells from ACh-induced death. Muscle Nerve, 2013. Copyright © 2013 Wiley Periodicals, Inc
Microglial Extracellular Vesicles as Modulators of Brain Microenvironment in Glioma
Full text linkMicroglial cells represent the resident immune elements of the central nervous system, where they exert constant monitoring and contribute to preserving neuronal activity and function. In the context of glioblastoma (GBM), a common type of tumor originating in the brain, microglial cells deeply modify their phenotype, lose their homeostatic functions, invade the tumoral mass and support the growth and further invasion of the tumoral cells into the surrounding brain parenchyma. These modifications are, at least in part, induced by bidirectional communication among microglial and tumoral cells through the release of soluble molecules and extracellular vesicles (EVs). EVs produced by GBM and microglial cells transfer different kinds of biological information to receiving cells, deeply modifying their phenotype and activity and could represent important diagnostic markers and therapeutic targets. Recent evidence demonstrates that in GBM, microglial-derived EVs contribute to the immune suppression of the tumor microenvironment (TME), thus favoring GBM immune escape. In this review, we report the current knowledge on EV formation, biogenesis, cargo and functions, with a focus on the effects of microglia-derived EVs in GBM. What clearly emerges from this analysis is that we are at the beginning of a full understanding of the complete picture of the biological effects of microglial-derived EVs and that further investigations using multidisciplinary approaches are necessary to validate their use in GBM diagnosis and therapy
Functional roles of the Ca2+-activated K+ channel, KCa3.1, in brain tumors
No full textGlioblastoma is the most aggressive and deadly brain tumor, with low disease-free period even after surgery and combined radio and chemotherapies. Among the factors contributing to rapid tumor growth in the brain are the elevated proliferation and invasion rate, and the ability to induce a local immunosuppressive environment. The intermediate-conductance Ca2+-activated K+ channel KCa3.1 is expressed on glioblastoma cells and in tumor-infiltrating cells. In tumor cells, the functional expression of KCa3.1 is important to modulate cell invasion and proliferation. In tumor infiltrating cells KCa3.1 activity is required to regulate their activation state. Interfering with KCa3.1 activity can be an adjuvant therapeutic approach in addition to classic chemotherapy, to counteract tumor growth and prolong patient's survival. In this mini-review we discuss the evidence of the functional roles of KCa3.1 channels in glioblastoma biolog
Fractalkine in the nervous system: neuroprotective or neurotoxic molecule?
No full textFractalkine (CX3CL1) is an intriguing chemokine that plays a central role in the nervous system. The expression of CX3CL1 on neurons and of its receptor CX3CR1 on microglia facilitates a privileged interaction, playing important roles in regulating the function and maturation of these cells. CX3CL1 is reported to have neuroprotective and anti-inflammatory activities in several experimental systems and animal models of disease, and its expression correlates with positive outcomes in human neuropathologies. However, a comparable amount of evidence shows that CX3CL1 sustains neuroinflammatory conditions and contributes to neurotoxicity. This review discusses the evidence in favor of the CX3CL1/CX3CR1 pair being neuroprotective and other evidence that it is neurotoxic. Our aim is to stimulate future research examining the molecular and cellular determinants responsible for this unique functional switch, which could be important for several neuropathologies
Cys residues are critical for chemokine receptor CXCR2 functional properties.
No full textAbstract
We examined the role of cysteine (Cys) residues present in chemokine receptor CXCR2 for proper surface expression, dimerization, signaling, and chemotaxis. To address this issue, serine or leucine residues were substituted for Cys, generating nine CXCR2 mutants transiently expressed in HEK cells. Single substitution of Cys residues present in the three extracellular loops (C119L, C196L, C286S) or in the seventh-transmembrane (TM) domain (C308L) abolished CXCL8 agonist binding, while no Cys substitution abolished surface receptor expression. We have previously demonstrated that CXCR2 dimerizes under reducing conditions, due to hydrophobic interactions that involve TM3 regions, and here we show that the dimer/monomer CXCR2 ratio drastically increases when analyzed under non-reducing conditions. We report that none of the Cys-deficient CXCR2 mutants abolishes receptor dimerization, demonstrating that Cys-Cys bonds are not the exclusive determinant of CXCR2 dimerization. Furthermore, both wt- and Cys-mutated CXCR2 dimers are expressed at the cell surface, indicating that receptor dimers are efficiently transferred at the plasma membrane. We also show that every Cys substitution in CXCR2, including those that still bind CXCL8, results in an impairment of receptor activity, analyzed as cell chemotaxis and intracellular signaling, suggesting that some structural requirement is likely fulfilled by Cys presence
Ligand-independent CXCR2 dimerization
No full textHomo- and hetero-oligomerization have been reported for several G protein-coupled receptors (GPCRs). The CXCR2 is a GPCR that is activated, among the others, by the chemokines CXCL8 (interleukin-8) and CXCL2 (growth-related gene product beta) to induce cell chemotaxis. We have investigated the oligomerization of CXCR2 receptors expressed in human embryonic kidney cells and generated a series of truncated mutants to determine whether they could negatively regulate the wild-type (wt) receptor functions. CXCR2 receptor oligomerization was also studied by coimmunoprecipitation of green fluorescent protein- and V5-tagged CXCR2. Truncated CXCR2 receptors retained their ability to form oligomers only if the region between the amino acids Ala-106 and Lys-163 was present. In contrast, all of the deletion mutants analyzed were able to form heterodimers with the wt CXCR2 receptor, albeit with different efficiency, competing for wt/wt dimer formation. The truncated CXCR2 mutants were not functional and, when coexpressed with wt CXCR2, interfered with receptor functions, impairing cell signaling and chemotaxis. When CXCR2 was expressed with the AMPA-type glutamate receptor GluR1, CXCR2 dimerization was again impaired in a dose-dependent way, and receptor functions were prejudiced. In contrast, CXCR1, a chemokine receptor that shares many similarities with CXCR2, did not dimerize alone or with CXCR2 and when coexpressed with CXCR2 did not impair receptor signaling and chemotaxis. The formation of CXCR2 dimers was also confirmed in cerebellar neuron cells. Taken together, we conclude from these studies that CXCR2 functions as a dimer and that truncated receptors negatively modulate receptor activities competing for the formation of wt/wt dimers
Functional properties of neurons derived from fetal mouseneurospheres are compatible with those of neuronal precursors in vivo.
No full textNeural stem cells can be propagated in culture as neurospheres, yielding neurons and glial cells upon differentiation. Although the neurosphere model is widely used, the functional properties of the neurosphere-derived neurons have been only partially characterized, and it is unclear whether repeated passaging alters their functional properties. In this study, we analyzed voltage- and transmitter-gated responses in neuron-like cells obtained by differentiating fetal mouse neurospheres at increasing passages in culture. We report that neurons fire overshooting action potentials in response to depolarizing currents up to passage 10 but loose this capability at later passages, as the density of voltage-gated Na+ and K+ currents decreases. In contrast, the immunoreactivity for the neuronal marker β-tubulin remains unaltered up to passage 21, indicating that this marker is not representative of cell function. In almost all neurons, γ-aminobutyric acid (GABA) evoked bicuculline-sensitive whole-cell currents, resulting from the activation of GABAA receptors, which appeared to be excitatory, insofar as the reversal potential of GABA-gated current was about −50 mV. Much smaller currents were elicited by the glutamatergic agonist AMPA, and only occasional responses to glycine were detected. In these functional aspects, neurosphere-derived neurons are similar to immature neurons differentiating in vivo. Therefore, at least for a limited number of passages in vitro, neurospheres provide an adequate model of in vivo neurogenesis
Chemokine CXCL8 modulates GluR1 phosphorylation
No full textThe chemokine interleukin 8/CXCL8 induces the phosphorylation of the GluR1 subunit of the AMPA-type glutamate receptor in neurons and transfected HEK cells, on both serine 845 (S845) and 831 (S831) residues. We previously described that CXCL8 receptor CXCR2 and GluR1 co-precipitate and that GluR1/CXCR2 co-expression both in HEK cells and neurons impairs CXCL8-induced cell migration. Here we show that replacement of S845 with Ala (A), but not with Glu (E), strongly reduces GluR1/CXCR2,2 interaction and abolishes the impairment of CXCL8-induced cell migration. Considered together our findings point to the phosphorylated state of S845GluR1 as a determinant of GluR1-CXCR2 physical coupling. (C) 2008 Elsevier B.V. All rights reserved
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