Publications by authors named "Carlos A Saura"

44 Publications

Tissue Clearing and Expansion Methods for Imaging Brain Pathology in Neurodegeneration: From Circuits to Synapses and Beyond.

Front Neurosci 2020 5;14:914. Epub 2020 Oct 5.

Institut de Neurociències, Departament de Bioquímica i Biologia Molecular, Facultat de Medicina, Universitat Autònoma de Barcelona, Barcelona, Spain.

Studying the structural alterations occurring during diseases of the nervous system requires imaging heterogeneous cell populations at the circuit, cellular and subcellular levels. Recent advancements in brain tissue clearing and expansion methods allow unprecedented detailed imaging of the nervous system through its entire scale, from circuits to synapses, including neurovascular and brain lymphatics elements. Here, we review the state-of-the-art of brain tissue clearing and expansion methods, mentioning their main advantages and limitations, and suggest their parallel implementation for circuits-to-synapses brain imaging using conventional (diffraction-limited) light microscopy -such as confocal, two-photon and light-sheet microscopy- to interrogate the cellular and molecular basis of neurodegenerative diseases. We discuss recent studies in which clearing and expansion methods have been successfully applied to study neuropathological processes in mouse models and postmortem human brain tissue. Volumetric imaging of cleared intact mouse brains and large human brain samples has allowed unbiased assessment of neuropathological hallmarks. In contrast, nanoscale imaging of expanded cells and brain tissue has been used to study the effect of protein aggregates on specific subcellular structures. Therefore, these approaches can be readily applied to study a wide range of brain processes and pathological mechanisms with cellular and subcellular resolution in a time- and cost-efficient manner. We consider that a broader implementation of these technologies is necessary to reveal the full landscape of cellular and molecular mechanisms underlying neurodegenerative diseases.
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http://dx.doi.org/10.3389/fnins.2020.00914DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7571329PMC
October 2020

Conventional and Non-Conventional Roles of Non-Muscle Myosin II-Actin in Neuronal Development and Degeneration.

Cells 2020 08 19;9(9). Epub 2020 Aug 19.

Institut de Neurociències, Department de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona, 08193 Barcelona, Spain.

Myosins are motor proteins that use chemical energy to produce mechanical forces driving actin cytoskeletal dynamics. In the brain, the conventional non-muscle myosin II (NMII) regulates actin filament cytoskeletal assembly and contractile forces during structural remodeling of axons and dendrites, contributing to morphology, polarization, and migration of neurons during brain development. NMII isoforms also participate in neurotransmission and synaptic plasticity by driving actin cytoskeletal dynamics during synaptic vesicle release and retrieval, and formation, maturation, and remodeling of dendritic spines. NMIIs are expressed differentially in cerebral non-neuronal cells, such as microglia, astrocytes, and endothelial cells, wherein they play key functions in inflammation, myelination, and repair. Besides major efforts to understand the physiological functions and regulatory mechanisms of NMIIs in the nervous system, their contributions to brain pathologies are still largely unclear. Nonetheless, genetic mutations or deregulation of NMII and its regulatory effectors are linked to autism, schizophrenia, intellectual disability, and neurodegeneration, indicating non-conventional roles of NMIIs in cellular mechanisms underlying neurodevelopmental and neurodegenerative disorders. Here, we summarize the emerging biological roles of NMIIs in the brain, and discuss how actomyosin signaling contributes to dysfunction of neurons and glial cells in the context of neurological disorders. This knowledge is relevant for a deep understanding of NMIIs on the pathogenesis and therapeutics of neuropsychiatric and neurodegenerative diseases.
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http://dx.doi.org/10.3390/cells9091926DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7566000PMC
August 2020

Adenosine A Receptor Antagonists Affects NMDA Glutamate Receptor Function. Potential to Address Neurodegeneration in Alzheimer's Disease.

Cells 2020 04 26;9(5). Epub 2020 Apr 26.

Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, Valderrebollo, 5, 28031 Madrid, Spain.

(1) Background. -methyl d-aspartate (NMDA) ionotropic glutamate receptor (NMDAR), which is one of the main targets to combat Alzheimer's disease (AD), is expressed in both neurons and glial cells. The aim of this paper was to assess whether the adenosine A receptor (AR), which is a target in neurodegeneration, may affect NMDAR functionality. (2) Methods. Immuno-histo/cytochemical, biophysical, biochemical and signaling assays were performed in a heterologous cell expression system and in primary cultures of neurons and microglia (resting and activated) from control and the APP transgenic mice. (3) Results. On the one hand, NMDA and A receptors were able to physically interact forming complexes, mainly in microglia. Furthermore, the amount of complexes was markedly enhanced in activated microglia. On the other hand, the interaction resulted in a novel functional entity that displayed a cross-antagonism, that could be useful to prevent the exacerbation of NMDAR function by using AR antagonists. Interestingly, the amount of complexes was markedly higher in the hippocampal cells from the APP than from the control mice. In neurons, the number of complexes was lesser, probably due to NMDAR not interacting with the AR. However, the activation of the AR receptors resulted in higher NMDAR functionality in neurons, probably by indirect mechanisms. (4) Conclusions. AR antagonists such as istradefylline, which is already approved for Parkinson's disease (Nouriast in Japan and Nourianz in the US), have potential to afford neuroprotection in AD in a synergistic-like fashion. i.e., via both neurons and microglia.
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http://dx.doi.org/10.3390/cells9051075DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7290564PMC
April 2020

Proteasomal-Mediated Degradation of AKAP150 Accompanies AMPAR Endocytosis during cLTD.

eNeuro 2020 Mar/Apr;7(2). Epub 2020 Apr 16.

Institut de Neurociències and Dpt. Bioquímica i Biologia Molecular, Universitat Autònoma de Barcelona, Cerdanyola del Vallès 08193, Spain

The number and function of synaptic AMPA receptors (AMPARs) tightly regulates excitatory synaptic transmission. Current evidence suggests that AMPARs are inserted into the postsynaptic membrane during long-term potentiation (LTP) and are removed from the membrane during long-term depression (LTD). Dephosphorylation of GluA1 at Ser-845 and enhanced endocytosis are critical events in the modulation of LTD. Moreover, changes in scaffold proteins from the postsynaptic density (PSD) could be also related to AMPAR regulation in LTD. In the present study we analyzed the effect of chemical LTD (cLTD) on A-kinase anchoring protein (AKAP)150 and AMPARs levels in mouse-cultured neurons. We show that cLTD induces AKAP150 protein degradation via proteasome, coinciding with GluA1 dephosphorylation at Ser-845 and endocytosis of GluA1-containing AMPARs. Pharmacological inhibition of proteasome activity, but not phosphatase calcineurin (CaN), reverted cLTD-induced AKAP150 protein degradation. Importantly, AKAP150 silencing induced dephosphorylation of GluA1 Ser-845 and GluA1-AMPARs endocytosis while AKAP150 overexpression blocked cLTD-mediated GluA1-AMPARs endocytosis. Our results provide direct evidence that cLTD-induced AKAP150 degradation by the proteasome contributes to synaptic AMPARs endocytosis.
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http://dx.doi.org/10.1523/ENEURO.0218-19.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7163082PMC
April 2020

Presenilin/γ-secretase-dependent EphA3 processing mediates axon elongation through non-muscle myosin IIA.

Elife 2019 10 2;8. Epub 2019 Oct 2.

Institut de Neurociències, Department de Bioquímica i Biologia Molecular, Facultat de Medicina, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona, Barcelona, Spain.

EphA/ephrin signaling regulates axon growth and guidance of neurons, but whether this process occurs also independently of ephrins is unclear. We show that presenilin-1 (PS1)/γ-secretase is required for axon growth in the developing mouse brain. PS1/γ-secretase mediates axon growth by inhibiting RhoA signaling and cleaving EphA3 independently of ligand to generate an intracellular domain (ICD) fragment that reverses axon defects in PS1/γ-secretase- and EphA3-deficient hippocampal neurons. Proteomic analysis revealed that EphA3 ICD binds to non-muscle myosin IIA (NMIIA) and increases its phosphorylation (Ser1943), which promotes NMIIA filament disassembly and cytoskeleton rearrangement. PS1/γ-secretase-deficient neurons show decreased phosphorylated NMIIA and NMIIA/actin colocalization. Moreover, pharmacological NMII inhibition reverses axon retraction in PS-deficient neurons suggesting that NMIIA mediates PS/EphA3-dependent axon elongation. In conclusion, PS/γ-secretase-dependent EphA3 cleavage mediates axon growth by regulating filament assembly through RhoA signaling and NMIIA, suggesting opposite roles of EphA3 on inhibiting (ligand-dependent) and promoting (receptor processing) axon growth in developing neurons.
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http://dx.doi.org/10.7554/eLife.43646DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6774734PMC
October 2019

Potentiation of cannabinoid signaling in microglia by adenosine A receptor antagonists.

Glia 2019 12 19;67(12):2410-2423. Epub 2019 Aug 19.

Centro de Investigación en Red, Enfermedades Neurodegenerativas (CiberNed), Instituto de Salud Carlos III, Madrid, Spain.

Neuroprotective M2-skewed microglia appear as promising to alter the course of neurodegenerative diseases and G protein-coupled receptors (GPCRs) are potential targets to achieve such microglial polarization. A common feature of adenosine A (A R) and cannabinoid CB (CB R) GPCRs in microglia is that their expression is upregulated in Alzheimer's disease (AD). On the one hand, CB R seems a target for neuroprotection, delaying neurodegenerative processes like those associated to AD or Parkinson's diseases. A R antagonists reduce amyloid burden and improve cognitive performance and memory in AD animal models. We here show a close interrelationship between these two receptors in microglia; they are able to physically interact and affect the signaling of each other, likely due to conformational changes within the A -CB receptor heteromer (A -CB Het). Particularly relevant is the upregulation of A -CB Het expression in samples from the APP , AD transgenic mice model. The most relevant finding, confirmed in both heterologous cells and in primary cultures of microglia, was that blockade of A receptors results in increased CB R-mediated signaling. This heteromer-specific feature suggests that A R antagonists would potentiate, via microglia, the neuroprotective action of endocannabinoids with implications for AD therapy.
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http://dx.doi.org/10.1002/glia.23694DOI Listing
December 2019

Altered microRNAs related to synaptic function as potential plasma biomarkers for Alzheimer's disease.

Alzheimers Res Ther 2019 05 15;11(1):46. Epub 2019 May 15.

Institut de Neurociències and Dpt. Bioquímica i Biología Molecular, Universitat Autònoma de Barcelona, 08193, Cerdanyola del Vallès, Spain.

Background: Several evidences suggest that failure of synaptic function occurs at preclinical stages of Alzheimer's disease (AD) preceding neuronal loss and the classical AD pathological hallmarks. Nowadays, there is an urgent need to identify reliable biomarkers that could be obtained with non-invasive methods to improve AD diagnosis at early stages. Here, we have examined plasma levels of a group of miRNAs related to synaptic proteins in a cohort composed of cognitive healthy controls (HC), mild cognitive impairment (MCI) and AD subjects.

Methods: Plasma and brain levels of miRNAs were analysed in two different cohorts including 38 HC, 26 MCI, 56 AD dementia patients and 27 frontotemporal dementia (FTD) patients. D'Agostino and Pearson and Shapiro-Wilk tests were used to evaluate data normality. miRNA levels between groups were compared using a two-sided nonparametric Mann-Whitney test and sensitivity and specificity was determined by receiver operating characteristic curve analysis.

Results: Significant upregulation of miR-92a-3p, miR-181c-5p and miR-210-3p was found in the plasma of both MCI and AD subjects. MCI patients that progress to AD showed higher plasma levels of these miRNAs. By contrast, no changes in miR-92a-3p, miR-181c-5p or miR-210-3p levels were observed in plasma obtained from a cohort of FTD.

Conclusion: Our study shows that plasma miR-92a-3p, miR-181c-5p and miR-210-3p constitute a specific molecular signature potentially useful as a potential biomarker for AD.
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http://dx.doi.org/10.1186/s13195-019-0501-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6521366PMC
May 2019

Synapse-to-Nucleus Signaling in Neurodegenerative and Neuropsychiatric Disorders.

Biol Psychiatry 2019 07 17;86(2):87-96. Epub 2019 Jan 17.

Institut de Neurociències, Department de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas, Universitat Autònoma de Barcelona, Barcelona, Spain. Electronic address:

Synapse-to-nucleus signaling is critical for converting signals received at synapses into transcriptional programs essential for cognition, memory, and emotion. This neuronal mechanism usually involves activity-dependent translocation of synaptonuclear factors from synapses to the nucleus resulting in regulation of transcriptional programs underlying synaptic plasticity. Acting as synapse-to-nucleus messengers, amyloid precursor protein intracellular domain associated-1 protein, cAMP response element binding protein (CREB)-regulated transcription coactivator-1, Jacob, nuclear factor kappa-light-chain-enhancer of activated B cells, RING finger protein 10, and SH3 and multiple ankyrin repeat domains 3 play essential roles in synapse remodeling and plasticity, which are considered the cellular basis of memory. Other synaptic proteins, such as extracellular signal-regulated kinase, calcium/calmodulin-dependent protein kinase II gamma, and CREB2, translocate from dendrites or cytosol to the nucleus upon synaptic activity, suggesting that they could contribute to synapse-to-nucleus signaling. Notably, some synaptonuclear factors converge on the transcription factor CREB, indicating that CREB signaling is a key hub mediating integration of synaptic signals into transcriptional programs required for neuronal function and plasticity. Although major efforts have been focused on identification and regulatory mechanisms of synaptonuclear factors, the relevance of synapse-to-nucleus communication in brain physiology and pathology is still unclear. Recent evidence, however, indicates that synaptonuclear factors are implicated in neuropsychiatric, neurodevelopmental, and neurodegenerative disorders, suggesting that uncoupling synaptic activity from nuclear signaling may prompt synapse pathology, contributing to a broad spectrum of brain disorders. This review summarizes current knowledge of synapse-to-nucleus signaling in neuron survival, synaptic function and plasticity, and memory. Finally, we discuss how altered synapse-to-nucleus signaling may lead to memory and emotional disturbances, which is relevant for clinical and therapeutic strategies in neurodegenerative and neuropsychiatric diseases.
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http://dx.doi.org/10.1016/j.biopsych.2019.01.006DOI Listing
July 2019

Decreased circulating ErbB4 ectodomain fragments as a read-out of impaired signaling function in amyotrophic lateral sclerosis.

Neurobiol Dis 2019 04 27;124:428-438. Epub 2018 Dec 27.

Instituto de Neurociencias de Alicante, Universidad Miguel Hernández-CSIC, Sant Joan d'Alacant, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain. Electronic address:

ErbB4 is a transmembrane receptor tyrosine kinase that binds to neuregulins to activate signaling. Proteolytic cleavage of ErbB4 results in release of soluble fragments of ErbB4 into the interstitial fluid. Disruption of the neuregulin-ErbB4 pathway has been suggested to be involved in the pathogenesis of amyotrophic lateral sclerosis (ALS). This study assesses whether soluble proteolytic fragments of the ErbB4 ectodomain (ecto-ErbB4) can be detected in cerebrospinal fluid (CSF) and plasma, and if the levels are altered in ALS. Immunoprecipitation combined with mass spectrometry or western blotting analyses confirmed the presence of ecto-ErbB4 in human CSF. Several anti-ErbB4-reactive bands, including a 55 kDa fragment, were detected in CSF. The bands were generated in the presence of neuregulin-1 (Nrg1) and were absent in plasma from ErbB4 knockout mice. Ecto-ErbB4 levels were decreased in CSF from ALS patients (n = 20) and ALS with concomitant frontotemporal dementia patients (n = 10), compared to age-matched controls (n = 13). A similar decrease was found for the short ecto-ErbB4 fragments in plasma of the same subjects. Likewise, the 55-kDa ecto-ErbB4 fragments were decreased in the plasma of the two transgenic mouse models of ALS (SOD1 and TDP-43). Intracellular ErbB4 fragments were decreased in the frontal cortex from SOD1 mice, indicating a reduction in Nrg-dependent induction of ErbB4 proteolytic processing, and suggesting impaired signaling. Accordingly, overexpression of Nrg1 induced by an adeno-associated viral vector increased the levels of the ecto-ErbB4 fragment in the SOD1 mice. We conclude that the determination of circulating ecto-ErbB4 fragments could be a tool to evaluate the impairment of the ErbB4 pathway and may be a useful biomarker in ALS.
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http://dx.doi.org/10.1016/j.nbd.2018.12.021DOI Listing
April 2019

-Methyl-D-Aspartate Receptor Link to the MAP Kinase Pathway in Cortical and Hippocampal Neurons and Microglia Is Dependent on Calcium Sensors and Is Blocked by α-Synuclein, Tau, and Phospho-Tau in Non-transgenic and Transgenic APP Mice.

Front Mol Neurosci 2018 28;11:273. Epub 2018 Aug 28.

Centro de Investigación en Red, Enfermedades Neurodegenerativas (CIBERNED), Instituto de Salud Carlos III, Madrid, Spain.

-methyl-D-aspartate receptors (NMDARs) respond to glutamate to allow the influx of calcium ions and the signaling to the mitogen-activated protein kinase (MAPK) cascade. Both MAPK- and Ca-mediated events are important for both neurotransmission and neural cell function and fate. Using a heterologous expression system, we demonstrate that NMDAR may interact with the EF-hand calcium-binding proteins calmodulin, calneuron-1, and NCS1 but not with caldendrin. NMDARs were present in primary cultures of both neurons and microglia from cortex and hippocampus. Calmodulin in microglia, and calmodulin and NCS1 in neurons, are necessary for NMDA-induced MAP kinase pathway activation. Remarkably, signaling to the MAP kinase pathway was blunted in primary cultures of cortical and hippocampal neurons and microglia from wild-type animals by proteins involved in neurodegenerative diseases: α-synuclein, Tau, and p-Tau. A similar blockade by pathogenic proteins was found using samples from the APP transgenic Alzheimer's disease model. Interestingly, a very marked increase in NMDAR-NCS1 complexes was identified in neurons and a marked increase of both NMDAR-NCS1 and NMDAR-CaM complexes was identified in microglia from the transgenic mice. The results show that α-synuclein, Tau, and p-Tau disrupt the signaling of NMDAR to the MAPK pathway and that calcium sensors are important for NMDAR function both in neurons and microglia. Finally, it should be noted that the expression of receptor-calcium sensor complexes, specially those involving NCS1, is altered in neural cells from APP mouse embryos/pups.
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http://dx.doi.org/10.3389/fnmol.2018.00273DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6127644PMC
August 2018

Molecular and functional interaction between GPR18 and cannabinoid CB G-protein-coupled receptors. Relevance in neurodegenerative diseases.

Biochem Pharmacol 2018 11 2;157:169-179. Epub 2018 Jun 2.

Molecular Neurobiology Laboratory, Department of Biochemistry and Molecular Biomedicine, University of Barcelona, Diagonal 643, 08028 Barcelona, Spain; Centro de Investigación en Red, Enfermedades Neurodegenerativas (CIBERNED), Instituto de Salud Carlos III, C/ Sinesio Delgado, 4, 28029 Madrid, Spain. Electronic address:

GPR18, still considered an orphan receptor, may respond to endocannabinoids, whose canonical receptors are CB and CB. GPR18 and CB receptors share a role in peripheral immune response regulation and are co-expressed in microglia, which are immunocompetent cells in the central nervous system (CNS). We aimed at identifying heteroreceptor complexes formed by GPR18 and CBR or CBR in resting and activated microglia. Receptor-receptor interaction was assessed using energy-transfer approaches, and receptor function by determining cAMP levels and ERK1/2 phosphorylation in heterologous cells and primary cultures of microglia. Heteroreceptor identification in primary cultures of microglia was achieved by in situ proximity ligation assays. Energy transfer results showed interaction of GPR18 with CBR but not with CBR. CB-GPR18 heteroreceptor complexes displayed particular functional properties (heteromer prints) often consisting of negative cross-talk (activation of one receptor reduces signaling arising from the partner receptor) and cross-antagonism (the response of one of the receptors is blocked by a selective antagonist of the partner receptor). Activated microglia showed the heteromer print (negative cross-talk and bidirectional cross-antagonism) and increased expression of CBR and GPR18. Due to the important role of CBR in neuroprotection, we further investigated heteroreceptor occurrence in primary cultures of microglia from transgenic mice overexpressing human APP, an Alzheimer's disease model. Microglial cells from transgenic mice showed the heteromer print and functional interactions that were similar to those found in cells from wild-type animals that were activated by treatment with lipopolysaccharide and interferon-γ. Our results suggest that GPR18 and its heteromers may play important roles in neurodegenerative processes.
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http://dx.doi.org/10.1016/j.bcp.2018.06.001DOI Listing
November 2018

Reversal of memory and neuropsychiatric symptoms and reduced tau pathology by selenium in 3xTg-AD mice.

Sci Rep 2018 04 24;8(1):6431. Epub 2018 Apr 24.

Institut de Neurociències, Departament de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona, Bellaterra, 08193, Spain.

Accumulation of amyloid-β plaques and tau contribute to the pathogenesis of Alzheimer's disease (AD), but it is unclear whether targeting tau pathology by antioxidants independently of amyloid-β causes beneficial effects on memory and neuropsychiatric symptoms. Selenium, an essential antioxidant element reduced in the aging brain, prevents development of neuropathology in AD transgenic mice at early disease stages. The therapeutic potential of selenium for ameliorating or reversing neuropsychiatric and cognitive behavioral symptoms at late AD stages is largely unknown. Here, we evaluated the effects of chronic dietary sodium selenate supplementation for 4 months in female 3xTg-AD mice at 12-14 months of age. Chronic sodium selenate treatment efficiently reversed hippocampal-dependent learning and memory impairments, and behavior- and neuropsychiatric-like symptoms in old female 3xTg-AD mice. Selenium significantly decreased the number of aggregated tau-positive neurons and astrogliosis, without globally affecting amyloid plaques, in the hippocampus of 3xTg-AD mice. These results indicate that selenium treatment reverses AD-like memory and neuropsychiatric symptoms by a mechanism involving reduction of aggregated tau and/or reactive astrocytes but not amyloid pathology. These results suggest that sodium selenate could be part of a combined therapeutic approach for the treatment of memory and neuropsychiatric symptoms in advanced AD stages.
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http://dx.doi.org/10.1038/s41598-018-24741-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5915484PMC
April 2018

CRTC1 mediates preferential transcription at neuronal activity-regulated CRE/TATA promoters.

Sci Rep 2017 12 21;7(1):18004. Epub 2017 Dec 21.

Institut de Neurociències, Department de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona, Barcelona, 08193, Spain.

Gene expression mediated by the transcription factor cAMP-responsive element-binding protein (CREB) is essential for a wide range of brain processes. The transcriptional coactivartor CREB-regulated transcription coactivator-1 (CRTC1) is required for efficient induction of CREB target genes during neuronal activity. However, the mechanisms regulating induction of specific CREB/CRTC1-dependent genes during neuronal activity remain largely unclear. Here, we investigated the molecular mechanisms regulating activity-dependent gene transcription upon activation of the CREB/CRTC1 signaling pathway in neurons. Depolarization and cAMP signals induce preferential transcription of activity-dependent genes containing promoters with proximal CRE/TATA sequences, such as c-fos, Dusp1, Nr4a1, Nr4a2 and Ptgs2, but not genes with proximal CRE/TATA-less promoters (e.g. Nr4a3, Presenilin-1 and Presenilin-2). Notably, biochemical and chromatin immunoprecipitation analyses reveal constitutive binding of CREB to target gene promoters in the absence of neuronal activity, whereas recruitment of CRTC1 to proximal CRE/TATA promoters depends on neuronal activity. Neuronal activity induces rapid CRTC1 dephosphorylation, nuclear translocation and binding to endogenous CREB. These results indicate that neuronal activity induces a preferential binding of CRTC1 to the transcriptional complex in CRE/TATA-containing promoters to engage activity-dependent transcription in neurons.
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http://dx.doi.org/10.1038/s41598-017-18215-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5740062PMC
December 2017

Proteolytic Processing of Neurexins by Presenilins Sustains Synaptic Vesicle Release.

J Neurosci 2018 01 11;38(4):901-917. Epub 2017 Dec 11.

Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/Consejo Superior de Investigaciones Científicas (CSIC)/Universidad de Sevilla, 41013 Sevilla, Spain,

Proteolytic processing of synaptic adhesion components can accommodate the function of synapses to activity-dependent changes. The adhesion system formed by neurexins (Nrxns) and neuroligins (Nlgns) bidirectionally orchestrate the function of presynaptic and postsynaptic terminals. Previous studies have shown that presenilins (PS), components of the gamma-secretase complex frequently mutated in familial Alzheimer's disease, clear from glutamatergic terminals the accumulation of Nrxn C-terminal fragments (Nrxn-CTF) generated by ectodomain shedding. Here, we characterized the synaptic consequences of the proteolytic processing of Nrxns in cultured hippocampal neurons from mice and rats of both sexes. We show that activation of presynaptic Nrxns with postsynaptic Nlgn1 or inhibition of ectodomain shedding in axonal Nrxn1-β increases presynaptic release at individual terminals, likely reflecting an increase in the number of functional release sites. Importantly, inactivation of PS inhibits presynaptic release downstream of Nrxn activation, leaving synaptic vesicle recruitment unaltered. Glutamate-receptor signaling initiates the activity-dependent generation of Nrxn-CTF, which accumulate at presynaptic terminals lacking PS function. The sole expression of Nrxn-CTF decreases presynaptic release and calcium flux, recapitulating the deficits due to loss of PS function. Our data indicate that inhibition of Nrxn processing by PS is deleterious to glutamatergic function. To gain insight into the role of presenilins (PS) in excitatory synaptic function, we address the relevance of the proteolytic processing of presynaptic neurexins (Nrxns) in glutamatergic differentiation. Using synaptic fluorescence probes in cultured hippocampal neurons, we report that trans-synaptic activation of Nrxns produces a robust increase in presynaptic calcium levels and neurotransmitter release at individual glutamatergic terminals by a mechanism that depends on normal PS activity. Abnormal accumulation of Nrxn C-terminal fragments resulting from impaired PS activity inhibits presynaptic calcium signal and neurotransmitter release, assigning synaptic defects to Nrxns as a specific PS substrate. These data may provide links into how loss of PS activity inhibits glutamatergic synaptic function in Alzheimer's disease patients.
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http://dx.doi.org/10.1523/JNEUROSCI.1357-17.2017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6596238PMC
January 2018

Emerging Roles of CREB-Regulated Transcription Coactivators in Brain Physiology and Pathology.

Trends Neurosci 2017 12 30;40(12):720-733. Epub 2017 Oct 30.

Center for Psychiatric Neuroscience and Service of Child and Adolescent Psychiatry, Department of Psychiatry, University Medical Center, University of Lausanne, Switzerland. Electronic address:

The brain has the ability to sense, coordinate, and respond to environmental changes through biological processes involving activity-dependent gene expression. cAMP-response element binding protein (CREB)-regulated transcription coactivators (CRTCs) have recently emerged as novel transcriptional regulators of essential biological functions, while their deregulation is linked to age-related human diseases. In the brain, CRTCs are unique signaling factors that act as sensors and integrators of hormonal, metabolic, and neural signals contributing to brain plasticity and brain-body communication. In this review, we focus on the regulatory mechanisms and functions of CRTCs in brain metabolism, lifespan, circadian rhythm, and synaptic mechanisms underlying memory and emotion. We also discuss how CRTCs deregulation in cognitive and emotional disorders may provide the basis for potential clinical and therapeutic applications in neurodegenerative and psychiatric diseases.
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http://dx.doi.org/10.1016/j.tins.2017.10.002DOI Listing
December 2017

Suspended Silicon Microphotodiodes for Electrochemical and Biological Applications.

Small 2017 11 25;13(41). Epub 2017 Sep 25.

Micro and Nano-Tools Group, Instituto de Microelectrónica de Barcelona IMB-CNM (CSIC), Campus UAB, Bellaterra, 08193, Spain.

Local electric stimulation of tissues and cells has gained importance as therapeutic alternative in the treatment of many diseases. These alternatives aim to deliver a less invasively stimuli in liquid media, making imperative the development of versatile micro- and nanoscale solutions for wireless actuation. Here, a simple microfabrication process to produce suspended silicon microphotodiodes that can be activated by visible light to generate local photocurrents in their surrounding medium is presented. Electrical characterization using electrical probes confirms their diode behavior. To demonstrate their electrochemical performance, an indirect test is implemented in solution through photoelectrochemical reactions controlled by a white-LED lamp. Furthermore, their effects on biological systems are observed in vitro using mouse primary neurons in which the suspended microphotodiodes are activated periodically with white-LED lamp, bringing out observable morphological changes in neuronal processes. The results demonstrate a simplified and cost-effective wireless tool for photovoltaic current generation in liquid media at the microscale.
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http://dx.doi.org/10.1002/smll.201701920DOI Listing
November 2017

Receptor-heteromer mediated regulation of endocannabinoid signaling in activated microglia. Role of CB and CB receptors and relevance for Alzheimer's disease and levodopa-induced dyskinesia.

Brain Behav Immun 2018 Jan 24;67:139-151. Epub 2017 Aug 24.

Molecular Neurobiology laboratory, Department of Biochemistry and Molecular Biomedicine, University of Barcelona, Diagonal 643, 08028 Barcelona, Spain; Centro de Investigación en Red, Enfermedades Neurodegenerativas (CIBERNED), Instituto de Salud Carlos III, C/ Sinesio Delgado, 4, 28029 Madrid, Spain. Electronic address:

Endocannabinoids are important regulators of neurotransmission and, acting on activated microglia, they are postulated as neuroprotective agents. Endocannabinoid action is mediated by CB and CB receptors, which may form heteromeric complexes (CB-CBHets) with unknown function in microglia. We aimed at establishing the expression and signaling properties of cannabinoid receptors in resting and LPS/IFN-γ-activated microglia. In activated microglia mRNA transcripts increased (2 fold for CB and circa 20 fold for CB), whereas receptor levels were similar for CB and markedly upregulated for CB; CB-CBHets were also upregulated. Unlike in resting cells, CB receptors became robustly coupled to G in activated cells, in which CB-CBHets mediated a potentiation effect. Hence, resting cells were refractory while activated cells were highly responsive to cannabinoids. Interestingly, similar results were obtained in cultures treated with ß-amyloid (Aß). Microglial activation markers were detected in the striatum of a Parkinson's disease (PD) model and, remarkably, in primary microglia cultures from the hippocampus of mutant β-amyloid precursor protein (APP) mice, a transgenic Alzheimer's disease (AD) model. Also of note was the similar cannabinoid receptor signaling found in primary cultures of microglia from APP and in cells from control animals activated using LPS plus IFN-γ. Expression of CB-CBHets was increased in the striatum from rats rendered dyskinetic by chronic levodopa treatment. In summary, our results showed sensitivity of activated microglial cells to cannabinoids, increased CB-CBHet expression in activated microglia and in microglia from the hippocampus of an AD model, and a correlation between levodopa-induced dyskinesia and striatal microglial activation in a PD model. Cannabinoid receptors and the CB-CB heteroreceptor complex in activated microglia have potential as targets in the treatment of neurodegenerative diseases.
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http://dx.doi.org/10.1016/j.bbi.2017.08.015DOI Listing
January 2018

CRTC1 Function During Memory Encoding Is Disrupted in Neurodegeneration.

Biol Psychiatry 2017 01 11;81(2):111-123. Epub 2016 Jul 11.

Institut de Neurociències, Department de Bioquímica i Biologia Molecular; and the; Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas, Universitat Autònoma de Barcelona, Barcelona, Spain. Electronic address:

Background: Associative memory impairment is an early clinical feature of dementia patients, but the molecular and cellular mechanisms underlying these deficits are largely unknown. In this study, we investigated the functional regulation of the cyclic adenosine monophosphate response element binding protein (CREB)-regulated transcription coactivator 1 (CRTC1) by associative learning in physiological and neurodegenerative conditions.

Methods: We evaluated the activation of CRTC1 in the hippocampus of control mice and mice lacking the Alzheimer's disease-linked presenilin genes (presenilin conditional double knockout [PS cDKO]) after one-trial contextual fear conditioning by using biochemical, immunohistochemical, and gene expression analyses. PS cDKO mice display classical features of neurodegeneration occurring in Alzheimer's disease including age-dependent cortical atrophy, neuron loss, dendritic degeneration, and memory deficits.

Results: Context-associative learning, but not single context or unconditioned stimuli, induces rapid dephosphorylation (Ser151) and translocation of CRTC1 from the cytosol/dendrites to the nucleus of hippocampal neurons in the mouse brain. Accordingly, context-associative learning induces differential CRTC1-dependent transcription of c-fos and the nuclear receptor subfamily 4 (Nr4a) genes Nr4a1-3 in the hippocampus through a mechanism that involves CRTC1 recruitment to CRE promoters. Deregulation of CRTC1 dephosphorylation, nuclear translocation, and transcriptional function are associated with long-term contextual memory deficits in PS cDKO mice. Importantly, CRTC1 gene therapy in the hippocampus ameliorates context memory and transcriptional deficits and dendritic degeneration despite ongoing cortical degeneration in this neurodegeneration mouse model.

Conclusions: These findings reveal a critical role of CRTC1 in the hippocampus during associative memory, and provide evidence that CRTC1 deregulation underlies memory deficits during neurodegeneration.
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http://dx.doi.org/10.1016/j.biopsych.2016.06.025DOI Listing
January 2017

Preservation of cell-survival mechanisms by the presenilin-1 K239N mutation may cause its milder clinical phenotype.

Neurobiol Aging 2016 10 15;46:169-79. Epub 2016 Jul 15.

Institut d'Investigacions Biomèdiques de Barcelona (IIBB), CSIC, Barcelona, Spain; Institut d'Investigació Biomèdica August Pi i Sunyer (IDIBAPS), Barcelona, Spain. Electronic address:

Presenilin 1 (PSEN1) mutations are the main cause of monogenic Alzheimer's disease. We studied the functional effects of the mutation K239N, which shows incomplete penetrance at the age of 65 years and compared it with the more aggressive mutation E120G. We engineered stable cell lines expressing human PSEN1 wild type or with K239N or E120G mutations. Both mutations induced dysfunction of γ-secretase in the processing of amyloid-β protein precursor, leading to an increase in the amyloid β42/amyloid β40 ratio. Analysis of homeostatic mechanisms showed that K239N induced lower basal and hydrogen peroxide induced intracellular levels of reactive oxygen species than E120G. Similarly, K239N induced lower vulnerability to apoptosis by hydrogen peroxide injury than E120G. Accordingly, the proapoptotic signaling pathways c-Jun NH2-terminal kinase and p38 mitogen-activated protein kinase maintained PSEN1-mediated negative regulation in K239N but not in E120G-bearing cells. Furthermore, the activation of the prosurvival signaling pathways mitogen-activated protein kinase/extracellular signal-regulated kinase and phosphoinositide 3-kinase/Akt was lower in E120G-bearing cells. Therefore, preservation of mechanisms regulating cell responses independent of amyloid-β protein precursor processing may account for the milder phenotype induced by the PSEN1 K239N mutation.
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http://dx.doi.org/10.1016/j.neurobiolaging.2016.07.002DOI Listing
October 2016

Gene expression parallels synaptic excitability and plasticity changes in Alzheimer's disease.

Front Cell Neurosci 2015 25;9:318. Epub 2015 Aug 25.

Institut de Neurociències, Departament de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona Barcelona, Spain.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by abnormal accumulation of β-amyloid and tau and synapse dysfunction in memory-related neural circuits. Pathological and functional changes in the medial temporal lobe, a region essential for explicit memory encoding, contribute to cognitive decline in AD. Surprisingly, functional imaging studies show increased activity of the hippocampus and associated cortical regions during memory tasks in presymptomatic and early AD stages, whereas brain activity declines as the disease progresses. These findings suggest an emerging scenario where early pathogenic events might increase neuronal excitability leading to enhanced brain activity before clinical manifestations of the disease, a stage that is followed by decreased brain activity as neurodegeneration progresses. The mechanisms linking pathology with synaptic excitability and plasticity changes leading to memory loss in AD remain largely unclear. Recent studies suggest that increased brain activity parallels enhanced expression of genes involved in synaptic transmission and plasticity in preclinical stages, whereas expression of synaptic and activity-dependent genes are reduced by the onset of pathological and cognitive symptoms. Here, we review recent evidences indicating a relationship between transcriptional deregulation of synaptic genes and neuronal activity and memory loss in AD and mouse models. These findings provide the basis for potential clinical applications of memory-related transcriptional programs and their regulatory mechanisms as novel biomarkers and therapeutic targets to restore brain function in AD and other cognitive disorders.
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http://dx.doi.org/10.3389/fncel.2015.00318DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4548151PMC
September 2015

Crtc1 activates a transcriptional program deregulated at early Alzheimer's disease-related stages.

J Neurosci 2014 Apr;34(17):5776-87

Institut de Neurociències, Department de Bioquímica i Biologia Molecular, Universitat Autònoma de Barcelona, Barcelona 08193, Spain, and Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED) and Institut de Neuropatologia, IDIBELL-Hospital Universitari de Bellvitge, Universitat de Barcelona, Barcelona 08907, Spain.

Cognitive decline is associated with gene expression changes in the brain, but the transcriptional mechanisms underlying memory impairments in cognitive disorders, such as Alzheimer's disease (AD), are largely unknown. Here, we aimed to elucidate relevant mechanisms responsible for transcriptional changes underlying early memory loss in AD by examining pathological, behavioral, and transcriptomic changes in control and mutant β-amyloid precursor protein (APPSw,Ind) transgenic mice during aging. Genome-wide transcriptome analysis using mouse microarrays revealed deregulation of a gene network related with neurotransmission, synaptic plasticity, and learning/memory in the hippocampus of APPSw,Ind mice after spatial memory training. Specifically, APPSw,Ind mice show changes on a cAMP-responsive element binding protein (CREB)-regulated transcriptional program dependent on the CREB-regulated transcription coactivator-1 (Crtc1). Interestingly, synaptic activity and spatial memory induces Crtc1 dephosphorylation (Ser151), nuclear translocation, and Crtc1-dependent transcription in the hippocampus, and these events are impaired in APPSw,Ind mice at early pathological and cognitive decline stages. CRTC1-dependent genes and CRTC1 levels are reduced in human hippocampus at intermediate Braak III/IV pathological stages. Importantly, adeno-associated viral-mediated Crtc1 overexpression in the hippocampus efficiently reverses Aβ-induced spatial learning and memory deficits by restoring a specific subset of Crtc1 target genes. Our results reveal a critical role of Crtc1-dependent transcription on spatial memory formation and provide the first evidence that targeting brain transcriptome reverses memory loss in AD.
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http://dx.doi.org/10.1523/JNEUROSCI.5288-13.2014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6608295PMC
April 2014

Ras protein activation is a key event in activity-dependent survival of cerebellar granule neurons.

J Biol Chem 2014 Mar 12;289(12):8462-72. Epub 2014 Feb 12.

From the Institut de Neurociencies and Department of Bioquímica i Biología Molecular, Universitat Autònoma de Barcelona, 08193 Cerdanyola del Vallès, Spain.

Neuronal activity promotes the survival of cerebellar granule neurons (CGNs) during the postnatal development of cerebellum. CGNs that fail to receive excitatory inputs will die by apoptosis. This process could be mimicked in culture by exposing CGNs to either a physiological concentration of KCl (5 mm or K5) plus N-methyl-d-aspartate (NMDA) or to 25 mm KCl (K25). We have previously described that a 24-h exposure to NMDA (100 μm) or K25 at 2 days in vitro induced long term survival of CGNs in K5 conditions. Here we have studied the molecular mechanisms activated at 2 days in vitro in these conditions. First we showed that NMDA or K25 addition promoted a rapid stimulation of PI3K and a biphasic phosphorylation on Ser-473 of Akt, a PI3K substrate. Interestingly, we demonstrated that only the first wave of Akt phosphorylation is necessary for the NMDA- and K25-mediated survival. Additionally, we detected that both NMDA and K25 increased ERK activity with a similar time-course. Moreover, our results showed that NMDA-mediated activation of the small G-protein Ras is necessary for PI3K/Akt pathway activation, whereas Rap1 was involved in NMDA phosphorylation of ERK. On the other hand, Ras, but not Rap1, mediates K25 activation of PI3K/Akt and MEK/ERK pathways. Because neuroprotection by NMDA or K25 is mediated by Ras (and not by Rap1) activation, we propose that Ras stimulation is a crucial event in NMDA- and K25-mediated survival of CGNs through the activation of PI3K/Akt and MEK/ERK pathways.
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http://dx.doi.org/10.1074/jbc.M113.536375DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961671PMC
March 2014

ApoER2 processing by presenilin-1 modulates reelin expression.

FASEB J 2014 Apr 16;28(4):1543-54. Epub 2013 Dec 16.

1Instituto de Neurociencias de Alicante, Universidad Miguel Hernández-CSIC, Av. Ramón y Cajal s/n, E-03550 Sant Joan d'Alacant, Spain.

The reelin signaling protein and its downstream components have been associated with synaptic plasticity and neurotransmission. The reelin signaling pathway begins with the binding of reelin to the transmembrane lipoprotein receptor apolipoprotein E receptor 2 (ApoER2), which in turns induces the sequential cleavage of ApoER2 by the sequential action of α- and γ-secretases. Using conditional-knockout mice of the catalytic component of the γ-secretase complex, presenilin 1 (PS1), we demonstrated increased brain ApoER2 and reelin protein and transcript levels, with no changes in the number of reelin-positive cells. Using the human SH-SY5Y neuroblastoma cell line, we showed that ApoER2 processing occurs in the presence of PS1, producing an intracellular ApoER2 C-terminal fragment. In addition, the pharmacologic inhibition of γ-secretase in SH-SY5Y cells led to increased reelin levels. Overexpression of ApoER2 decreased reelin mRNA levels in these cells. A luciferase reporter gene assay and nuclear fractionation confirmed that increased amounts of intracellular fragment of ApoER2 suppressed reelin expression at a transcriptional level. Chromatin immunoprecipitation experiments corroborated that the intracellular fragment of ApoER2 bound to the RELN promoter region. Our study suggests that PS1/γ-secretase-dependent processing of the reelin receptor ApoER2 inhibits reelin expression and may regulate its signaling.
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http://dx.doi.org/10.1096/fj.13-239350DOI Listing
April 2014

CSF Presenilin-1 complexes are increased in Alzheimer's disease.

Acta Neuropathol Commun 2013 Aug 6;1:46. Epub 2013 Aug 6.

Instituto de Neurociencias de Alicante, Universidad Miguel Hernández-CSIC, Av, Ramón y Cajal s/n, Sant Joan d'Alacant E-03550, Spain.

Background: Presenilin-1 (PS1) is the active component of the amyloid precursor protein cleaving γ-secretase complex. PS1 protein is a transmembrane protein containing multiple hydrophobic regions which presence in cerebrospinal fluid (CSF) has not been measured to date. This study assesses whether PS1 and other components of the γ-secretase complex are present in CSF.

Results: Here, we show that PS1 is present in ventricular post-mortem and lumbar ante-mortem CSF, and plasma as 100-150-kDa hetero-complexes containing both the N- and C-terminal fragments (NTF and CTF) of the protein. Immunoprecipitation and immunoblotting with different antibodies confirmed the identity of the PS1 species. The γ-secretase components, APH-1 (anterior pharynx-defective 1) and PEN-2 (presenilin enhancer 2), as well as presenilin-2 (PS2) fragments, co-exist within these CSF complexes, while nicastrin is not detected. These CSF-PS1 complexes differ from active γ-secretase membrane-complexes, and may represent nonspecific aggregation of the PS1 protein. Levels of PS1 complexes are increased in CSF samples from autopsy-confirmed Alzheimer's disease (AD) cases and were found to be more stable than complexes in CSF from control subjects. Despite similar levels of total PS1 in CSF from probable AD patients and cognitively normal subjects, an increased proportion of highly stable PS1 complexes were observed in AD CSF.

Conclusions: Our data suggest that fragments of the PS1 protein present in CSF as complexes may be useful as a biomarker for AD.
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http://dx.doi.org/10.1186/2051-5960-1-46DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3893612PMC
August 2013

X-linked inhibitor of apoptosis protein negatively regulates neuronal differentiation through interaction with cRAF and Trk.

Sci Rep 2013 ;3:2397

Institut de Neurociències and Dpt. Bioquímica and Biología Molecular, Universitat Autònoma de Barcelona, 08193 Cerdanyola del Vallès, Spain.

X-linked Inhibitor of apoptosis protein (XIAP) has been classically identified as a cell death regulator. Here, we demonstrate a novel function of XIAP as a regulator of neurite outgrowth in neuronal cells. In PC12 cells, XIAP overexpression prevents NGF-induced neuronal differentiation, whereas NGF treatment induces a reduction of endogenous XIAP levels concomitant with the induction of neuronal differentiation. Accordingly, downregulation of endogenous XIAP protein levels strongly increases neurite outgrowth in PC12 cells as well as axonal and dendritic length in primary cortical neurons. The effects of XIAP are mediated by the mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinases (ERKs) pathway since blocking this pathway completely prevents the neuritogenesis mediated by XIAP downregulation. In addition, we found that XIAP binds to cRaf and Trk receptors. Our results demonstrate that XIAP plays a new role as a negative regulator of neurotrophin-induced neurite outgrowth and neuronal differentiation in developing neurons.
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http://dx.doi.org/10.1038/srep02397DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3739015PMC
February 2014

ATP and noradrenaline activate CREB in astrocytes via noncanonical Ca(2+) and cyclic AMP independent pathways.

Glia 2012 Sep 16;60(9):1330-44. Epub 2012 May 16.

Institut de Neurociències, Universitat Autònoma de Barcelona, Spain.

In neurons, it is well established that CREB contributes to learning and memory by orchestrating the translation of experience into the activity-dependent (i.e., driven by neurotransmitters) transcription of plasticity-related genes. The activity-dependent CREB-triggered transcription requires the concerted action of cyclic AMP/protein kinase A and Ca(2+) /calcineurin via the CREB-regulated transcription co-activator (CRTC). It is not known, however, whether a comparable molecular sequence occurs in astrocytes, despite the unquestionable contribution of these cells to brain plasticity. Here we sought to determine whether and how ATP and noradrenaline cause CREB-dependent transcription in rat cortical astrocyte cultures. Both transmitters induced CREB phosphorylation (Western Blots), CREB-dependent transcription (CRE-luciferase reporter assays), and the transcription of Bdnf, a canonical regulator of synaptic plasticity (quantitative RT-PCR). We indentified a Ca(2+) and diacylglycerol-independent protein kinase C at the uppermost position of the cascade leading to CREB-dependent transcription. Notably, CREB-dependent transcription was partially dependent on ERK1/2 and CRTC, but independent of cyclic AMP/protein kinase A or Ca(2+) /calcineurin. We conclude that ATP and noradrenaline activate CREB-dependent transcription in cortical astrocytes via an atypical protein kinase C. It is of relevance that the signaling involved be starkly different to the one described in neurons since there is no convergence of Ca(2+) and cyclic AMP-dependent pathways on CRTC, which, moreover, exerts a modulatory rather than a central role. Our data thus point to the existence of an alternative, non-neuronal, glia-based role of CREB in plasticity.
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http://dx.doi.org/10.1002/glia.22352DOI Listing
September 2012

Nurr1 protein is required for N-methyl-D-aspartic acid (NMDA) receptor-mediated neuronal survival.

J Biol Chem 2012 Mar 31;287(14):11351-62. Epub 2012 Jan 31.

Institut de Neurociencies and Departament de Bioquímica i Biología Molecular, Universitat Autònoma de Barcelona, 08193 Barcelona, Spain.

NMDA receptor (NMDAR) stimulation promotes neuronal survival during brain development. Cerebellar granule cells (CGCs) need NMDAR stimulation to survive and develop. These neurons differentiate and mature during its migration from the external granular layer to the internal granular layer, and lack of excitatory inputs triggers their apoptotic death. It is possible to mimic this process in vitro by culturing CGCs in low KCl concentrations (5 mm) in the presence or absence of NMDA. Using this experimental approach, we have obtained whole genome expression profiles after 3 and 8 h of NMDA addition to identify genes involved in NMDA-mediated survival of CGCs. One of the identified genes was Nurr1, a member of the orphan nuclear receptor subfamily Nr4a. Our results report a direct regulation of Nurr1 by CREB after NMDAR stimulation. ChIP assay confirmed CREB binding to Nurr1 promoter, whereas CREB shRNA blocked NMDA-mediated increase in Nurr1 expression. Moreover, we show that Nurr1 is important for NMDAR survival effect. We show that Nurr1 binds to Bdnf promoter IV and that silencing Nurr1 by shRNA leads to a decrease in brain-derived neurotrophic factor (BDNF) protein levels and a reduction of NMDA neuroprotective effect. Also, we report that Nurr1 and BDNF show a similar expression pattern during postnatal cerebellar development. Thus, we conclude that Nurr1 is a downstream target of CREB and that it is responsible for the NMDA-mediated increase in BDNF, which is necessary for the NMDA-mediated prosurvival effect on neurons.
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http://dx.doi.org/10.1074/jbc.M111.272427DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3322860PMC
March 2012

CREB-regulated transcription coactivator 1-dependent transcription in Alzheimer's disease mice.

Authors:
Carlos A Saura

Neurodegener Dis 2012 6;10(1-4):250-2. Epub 2012 Jan 6.

Institut de Neurociències, Departament Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas, Universitat Autònoma de Barcelona, Bellaterra, Spain.

Background/aims: Long-term memory requires fine-tuning regulation of gene expression in specific neural circuits of the brain. Transcriptional regulation of gene programs is a key mechanism for memory storage and its deregulation may contribute to synaptic and cognitive dysfunction in memory disorders. The molecular mechanisms underlying changes on activity-dependent gene expression in Alzheimer's disease (AD) are largely unknown.

Methods: We analyzed the expression of activity-dependent genes regulated by the cAMP response element binding protein (CREB) and activation of CREB and its coactivator CREB-regulated transcription coactivator 1 (CRTC1) in control and mutant β-amyloid precursor protein (APP(Sw,Ind); Swedish and Indiana mutations) transgenic mice.

Results: Gene expression analyses revealed specific downregulation of a subset of well-known activity-induced CREB-dependent genes, including c-fos, Bdnf and Nr4a2, in the hippocampus of memory-impaired APP(Sw,Ind) transgenic mice. Activity-dependent CREB transcription induced by calcium/cAMP signals is disrupted through a mechanism involving deregulation of calcium/calcineurin-mediated dephosphorylation and activation of CRTC1. Expression of CRTC1 and pharmacological activation of L-type voltage-gated calcium channels reverse the deficits in CRTC1-mediated transcription in APP(Sw,Ind) neurons.

Conclusion: Our results suggest that CRTC1 dysfunction caused by Aβ accumulation underlies changes in gene expression required for hippocampal-dependent memory in AD transgenic mice.
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http://dx.doi.org/10.1159/000333341DOI Listing
August 2012

Soluble oligomers of amyloid-β peptide disrupt membrane trafficking of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor contributing to early synapse dysfunction.

J Biol Chem 2011 Aug 10;286(31):27311-21. Epub 2011 Jun 10.

Institut de Neurociencies and Departament de Bioquímica i Biología Molecular, Universitat Autònoma de Barcelona, 08193 Cerdanyola del Vallès, Barcelona.

β-Amyloid (Aβ), a peptide generated from the amyloid precursor protein, is widely believed to underlie the pathophysiology of Alzheimer disease (AD). Emerging evidences suggest that soluble Aβ oligomers adversely affect synaptic function, leading to cognitive failure associated with AD. The Aβ-induced synaptic dysfunction has been attributed to the synaptic removal of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors (AMPARs). However, the molecular mechanisms underlying the loss of AMPAR induced by Aβ at synapses are largely unknown. In this study we have examined the effect of Aβ oligomers on phosphorylated GluA1 at serine 845, a residue that plays an essential role in the trafficking of AMPARs toward extrasynaptic sites and the subsequent delivery to synapses during synaptic plasticity events. We found that Aβ oligomers reduce basal levels of Ser-845 phosphorylation and surface expression of AMPARs affecting AMPAR subunit composition. Aβ-induced GluA1 dephosphorylation and reduced receptor surface levels are mediated by an increase in calcium influx into neurons through ionotropic glutamate receptors and activation of the calcium-dependent phosphatase calcineurin. Moreover, Aβ oligomers block the extrasynaptic delivery of AMPARs induced by chemical synaptic potentiation. In addition, reduced levels of total and phosphorylated GluA1 are associated with initial spatial memory deficits in a transgenic mouse model of AD. These findings indicate that Aβ oligomers could act as a synaptic depressor affecting the mechanisms involved in the targeting of AMPARs to the synapses during early stages of the disease.
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http://dx.doi.org/10.1074/jbc.M111.227504DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3149325PMC
August 2011

Presenilin/γ-secretase regulates neurexin processing at synapses.

PLoS One 2011 Apr 29;6(4):e19430. Epub 2011 Apr 29.

Institut de Neurociències, Departament de Bioquímica i Biologia Molecular, Centro de Investigación Biomédica en Red Enfermedades Neurodegenerativas (CIBERNED), Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain.

Neurexins are a large family of neuronal plasma membrane proteins, which function as trans-synaptic receptors during synaptic differentiation. The binding of presynaptic neurexins to postsynaptic partners, such as neuroligins, has been proposed to participate in a signaling pathway that regulates synapse formation/stabilization. The identification of mutations in neurexin genes associated with autism and mental retardation suggests that dysfunction of neurexins may underlie synaptic defects associated with brain disorders. However, the mechanisms that regulate neurexin function at synapses are still unclear. Here, we show that neurexins are proteolytically processed by presenilins (PS), the catalytic components of the γ-secretase complex that mediates the intramembraneous cleavage of several type I membrane proteins. Inhibition of PS/γ-secretase by using pharmacological and genetic approaches induces a drastic accumulation of neurexin C-terminal fragments (CTFs) in cultured rat hippocampal neurons and mouse brain. Neurexin-CTFs accumulate mainly at the presynaptic terminals of PS conditional double knockout (PS cDKO) mice lacking both PS genes in glutamatergic neurons of the forebrain. The fact that loss of PS function enhances neurexin accumulation at glutamatergic terminals mediated by neuroligin-1 suggests that PS regulate the processing of neurexins at glutamatergic synapses. Interestingly, presenilin 1 (PS1) is recruited to glutamatergic terminals mediated by neuroligin-1, thus concentrating PS1 at terminals containing β-neurexins. Furthermore, familial Alzheimer's disease (FAD)-linked PS1 mutations differentially affect β-neurexin-1 processing. Expression of PS1 M146L and PS1 H163R mutants in PS-/- cells rescues the processing of β-neurexin-1, whereas PS1 C410Y and PS1 ΔE9 fail to rescue the processing defect. These results suggest that PS regulate the synaptic function and processing of neurexins at glutamatergic synapses, and that impaired neurexin processing by PS may play a role in FAD.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0019430PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3084856PMC
April 2011