Disruption of striatal glutamatergic transmission induced by mutant huntingtin involves remodelling of both postsynaptic density and NMDA receptor signalling

We study the striatal susceptibility to NMDA receptor (NMDAR)-mediated injury of two Huntington’s disease (HD) transgenic mice:R6/1 and R6/1:BDNF+/−. We found that R6/1:BDNF+/− mice – whichexpress reduced levels of BDNF – were more resistant than R6/1 miceto intrastriatal injection of quinolinate. This increased resistance isrelated to a differential reduction in expression of NMDAR scaffoldingproteins, MAGUKs (PSD-95, PSD-93, SAP-102 and SAP-97) but notto altered levels or synaptic location of NMDAR. A robust reorganizationof postsynaptic density (PSD) was detected in HD transgenicmice, shown by a switch of PSD-93 by PSD-95 in PSD. Furthermore,NMDAR signaling pathways were affected by different BDNF levels inHD mice; we found a reduction of synaptic αCaMKII (but not ofnNOS) in R6/1:BDNF+/− compared to R6/1 mice. The specific regulationof MAGUKs and αCaMKII in striatal neurons may reflect aprotective mechanism against expression of mutant huntingtin exon-1

Altered m6A RNA methylation contributes to hippocampal memory deficits in Huntington's disease mice.

N6-methyladenosine (m6A) regulates many aspects of RNA metabolism and is involved in learning and memory processes. Yet, the impact of a dysregulation of post-transcriptional m6A editing on synaptic impairments in neurodegenerative disorders remains unknown. Here we investigated the m6A methylation pattern in the hippocampus of Huntington's disease (HD) mice and the potential role of the m6A RNA modification in HD cognitive symptomatology. m6A modifications were evaluated in HD mice subjected to a hippocampal cognitive training task through m6A immunoprecipitation sequencing (MeRIP-seq) and the relative levels of m6A-modifying proteins (FTO and METTL14) by subcellular fractionation and Western blot analysis. Stereotaxic CA1 hippocampal delivery of AAV-shFTO was performed to investigate the effect of RNA m6A dysregulation in HD memory deficits. Our results reveal a m6A hypermethylation in relevant HD and synaptic related genes in the hippocampal transcriptome of Hdh+/Q111 mice. Conversely, m6A is aberrantly regulated in an experience-dependent manner in the HD hippocampus leading to demethylation of important components of synapse organization. Notably, the levels of RNA demethylase (FTO) and methyltransferase (METTL14) were modulated after training in the hippocampus of WT mice but not in Hdh+/Q111 mice. Finally, inhibition of FTO expression in the hippocampal CA1 region restored memory disturbances in symptomatic Hdh+/Q111 mice. Altogether, our results suggest that a differential RNA methylation landscape contributes to HD cognitive symptoms and uncover a role of m6A as a novel hallmark of HD

Lack of Helios during neural development induces adult schizophrenia-like behaviors associated with aberrant levels of the TRIF-recruiter protein WDFY1

The role of the WDFY1 protein has been studied as a TLR3/4 scaffold/recruiting protein in the immune system and in different oncogenic conditions. However, its function in brain remains poorly understood. We have found that in mice devoid of Helios (He−/− mice), a transcription factor specifically expressed during the development of the immune cells and the central nervous system, there is a permanent and sustained increase of Wdfy1 gene expression in the striatum and hippocampus. Interestingly, we observed that WDFY1 protein levels were also increased in the hippocampus and dorsolateral prefrontal cortex of schizophrenic patients, but not in the hippocampus of Alzheimer's disease patients with an associated psychotic disorder. Accordingly, young He−/− mice displayed several schizophrenic-like behaviors related to dysfunctions in the striatum and hippocampus. These changes were associated with an increase in spine density in medium spiny neurons (MSNs) and with a decrease in the number and size of PSD-95-positive clusters in the stratum radiatum of the CA1. Moreover, these alterations in structural synaptic plasticity were associated with a strong reduction of neuronal NF-κB in the pyramidal layer of the CA1 in He−/− mice. Altogether, our data indicate that alterations involving the molecular axis Helios-WDFY1 in neurons during the development of core brain regions could be relevant for the pathophysiology of neuropsychiatric disorders such as schizophrenia

Helios expression coordinates the development of a subset of striatopallidal medium spiny neurons

Here we unravel the mechanism of action of Helios (He) during the development of striatal medium spiny neurons (MSNs). He regulates the second wave of striatal neurogenesis involved in the generation of striatopallidal neurons that express dopamine 2 receptor (D2R) and enkephalin (ENK). To exert this effect He is expressed in neural progenitor cells (NPCs) retaining them into the G1/G0 phase of the cell cycle. Thus, the lack of He produces an increase of S-phase entry and S-phase length of NPCs which in turn impairs striatal neurogenesis and produces an accumulation of the number of cycling NPCs in the germinal zone (GZ) that end up dying at postnatal stages. Therefore, He-/- mice show a reduction in the number of Dorso-Medial Striatal MSNs in the adulthood that produces deficits in motor skills acquisition. In addition, overexpression of He in NPCs induce DARPP32 phenotype when transplanted in mouse striatum.Present findings demonstrate that He is involved in the correct development of a subset of striatopallidal MSNs and reveal new cellular mechanisms for neuronal development

Disease-specific phenotypes in dopamine neurons from human iPS-based models of genetic and sporadic Parkinson's disease

Induced pluripotent stem cells (iPSC) offer an unprecedented opportunity to model human disease in relevant cell types, but it is unclear whether they could successfully model age-related diseases such as Parkinson's disease (PD). Here, we generated iPSC lines from seven patients with idiopathic PD (ID-PD), four patients with familial PD associated to the G2019S mutation in the Leucine-Rich Repeat Kinase 2 (LRRK2) gene (LRRK2-PD) and four age- and sex-matched healthy individuals (Ctrl). Over long-time culture, dopaminergic neurons (DAn) differentiated from either ID-PD- or LRRK2-PD-iPSC showed morphological alterations, including reduced numbers of neurites and neurite arborization, as well as accumulation of autophagic vacuoles, which were not evident in DAn differentiated from Ctrl-iPSC. Further induction of autophagy and/or inhibition of lysosomal proteolysis greatly exacerbated the DAn morphological alterations, indicating autophagic compromise in DAn from ID-PD- and LRRK2-PD-iPSC, which we demonstrate occurs at the level of autophagosome clearance. Our study provides an iPSC-based in vitro model that captures the patients' genetic complexity and allows investigation of the pathogenesis of both sporadic and familial PD cases in a disease-relevant cell type

Helios modulates the maturation of a CA1 neuronal subpopulation required for spatial memory formation

Currently, molecular, electrophysiological and structural studies delineate several neural subtypes in the hippocampus. However, the precise developmental mechanisms that lead to this diversity are still unknown. Here we show that alterations in a concrete hippocampal neuronal subpopulation during development specifically affect hippocampal-dependent spatial memory. We observed that the genetic deletion of the transcription factor Helios in mice, which is specifically expressed in developing hippocampal calbindin-positive CA1 pyramidal neurons (CB-CA1-PNs), induces adult alterations affecting spatial memory. In the same mice, CA3-CA1 synaptic plasticity and spine density and morphology in adult CB-CA1-PNs were severely compromised. RNAseq experiments in developing hippocampus identified an aberrant increase on the Visinin-like protein 1 (VSNL1) expression in the hippocampi devoid of Helios. This aberrant increase on VSNL1 levels was localized in the CB-CA1-PNs. Normalization of VSNL1 levels in CB-CA1-PNs devoid of Helios rescued their spine loss in vitro. Our study identifies a novel and specific developmental molecular pathway involved in the maturation and function of a CA1 pyramidal neuronal subtype

Estudi de l’expressió i funció del gen SIRT3 en el teixit adipós marró

[cat] SIRT3, membre de la família d’histona desacetilases anomenades sirtuïnes, es troba principalment al mitocondri de teixits amb alta capacitat oxidativa com el fetge, el múscul, el cor o teixit adipós marró i s’ha suggerit que podria jugar un paper en el metabolisme energètic. Per tal d’aclarir el seu rol en la termogènesi, es van analitzar les alteracions en ratolins SIRT3 -/- en dues condicions de termogènesi augmentada: la transició fetal-neonatal i l’exposició crònica al fred en ratolins adults, així com es va estudiar l’efecte del dejuni en adults SIRT3 -/- , una situació en la qual la termogènesi es troba inhibida. Es van avaluar els possibles trastorns metabòlics i es va procedir a un anàlisi transcriptòmic de l’expressió de gens marcadors de diverses vies metabòliques en el teixit adipós marró i altres teixits dels ratolins SIRT3 -/-. Els resultats obtinguts indiquen alteracions significatives en el període perinatal com a conseqüència de la manca de SIRT3, però no en l’etapa adulta. Concretament s’observa una expressió alterada de gens específics de la termogènesi (uncoupling protein-1, 5’-deiodinasa, PRDM16) al teixit adipós marró en l’etapa perinatal així com de gens implicats en l’oxidació d’àcids grassos (enoil-CoA, hidratasa/3-hidroxiacil CoA deshidrogenasa, acil-CoA oxidasa) al fetge durant l’etapa perinatal. Aquests resultats posen de manifest el potencial paper de SIRT3 en aquestes vies metabòliques i evidencien l’existència de mecanismes homeostàtics durant el desenvolupament post-natal capaços de compensar l’absència de SIRT3 en l’animal adult. D’altra banda, mitjançant l’ús d’adipòcits SIRT3 -/-, vam determinat que SIRT3 és necessària per una resposta apropiada de les cèl•lules a l’activació noradrenèrgica mediada per AMPc de l’expressió de gens termogènics del teixit adipós marró. El co-activador transcripcional PGC1α (peroxisome proliferator-activated receptor-γ coactivator-1α) indueix l’expressió de SIRT3 en adipòcits blancs i fibroblasts embrionaris com a part de la inducció global del patró d’expressió gènica específic del teixit adipós marró. A les cèl•lules sense SIRT3, PGC1α no pot induir completament l’expressió de gens termogènics específics de l’adipòcit marró. PGC1α activa la transcripció del gen SIRT3 a través de la coactivació del receptor nuclear orfe ERRα (estrogen-related receptor-α) el qual s’uneix a la regió proximal del promotor del gen SIRT3. Assajos de pèrdua de funció d’ERRα, indiquen que aquest és necessari per a la completa inducció de la l’expressió del gen SIRT3 en resposta a PGC1α. Aquests resultats indiquen que PGC1α controla l’expressió de SIRT3 i que aquesta acció és un component essencial del mecanisme global a través del qual PGC1α indueix la completa adquisició del fenotip d’adipòcit marró diferenciat.[eng] SIRT3 is a member of the sirtuin family of deacetylases present mainly in mitochondria from tissues such as brown adipose tissue, liver, muscle and heart. To gain insight in the role of SIRT3 in thermogenesis, we determined the alterations in mice with targeted invalidation of the SIRT3 gene (SIRT3-/- mice) in two conditions of enhanced thermogenesis: the fetal-to-neonatal transition and chronic exposure of adult mice to cold, an in fasting of adult mice, a condition of depressed thermogenesis. Assessment of overall metabolic disturbances and a transcritomic analysis of the expression of marker genes of distinct metabolic pathways were performed in brown fat and other tissues from SIRT3-/- mice in these conditions. Results indicated significant alterations in the perinatal period but not in adulthood. Impaired expression of specific genes of thermogenesis in perinatal brown fat, and of genes encoding components of the fatty acid oxidation machinery in perinatal liver were observed in SIRT3-/- mice. Results highlight the potential role of SIRT3 in these pathways and evidence the appearance of homeostatic mechanisms during post-natal development capable to compensate for the absence of SIRT3 in the adulthood. Sirt3 is expressed in association with brown adipocyte differentiation. Using Sirt3-null brown adipocytes, we determined that Sirt3 is required for an appropriate responsiveness of cells to noradrenergic, cAMP-mediated activation of the expression of brown adipose tissue thermogenic genes. The transcriptional coactivator Pgc-1α (peroxisome proliferator-activated receptor-γ coactivator-1 α) induced Sirt3 gene expression in white adipocytes and embryonic fibroblasts as part of its overall induction of a brown adipose tissue-specific pattern of gene expression. In cells lacking Sirt3, Pgc-1α failed to fully induce the expression of brown fat-specific thermogenic genes. Pgc-1α activates Sirt3 gene transcription through coactivation of the orphan nuclear receptor Err (estrogen-related receptor)- α, which bound the proximal Sirt3 gene promoter region. Errα knockdown assays indicated that Errα is required for full induction of Sirt3 gene expression in response to Pgc-1 α. The present study indicates that Pgc-1α controls Sirt3 gene expression and this action is an essential component of the overall mechanisms by which Pgc-1α induces the full acquisition of a brown adipocyte differentiated phenotype

Unraveling the spatiotemporal distribution of VPS13A in the mouse brain

Loss‐of‐function mutations in the human vacuolar protein sorting the 13 homolog A (VPS13A) gene cause Chorea‐acanthocytosis (ChAc), with selective degeneration of the striatum as the main neuropathologic feature. Very little is known about the VPS13A expression in the brain. The main objective of this work was to assess, for the first time, the spatiotemporal distribution of VPS13A in the mouse brain. We found VPS13A expression present in neurons already in the em‐bryonic stage, with stable levels until adulthood. VPS13A mRNA and protein distributions were similar in the adult mouse brain. We found a widespread VPS13A distribution, with the strongest expression profiles in the pons, hippocampus, and cerebellum. Interestingly, expression was weak in the basal ganglia. VPS13A staining was positive in glutamatergic, GABAergic, and cholinergic neurons, but rarely in glial cells. At the cellular level, VPS13A was mainly located in the soma and neurites, co‐localizing with both the endoplasmic reticulum and mitochondria. However, it was not enriched in dendritic spines or the synaptosomal fraction of cortical neurons. In vivo pharma‐cological modulation of the glutamatergic, dopaminergic or cholinergic systems did not modulate VPS13A concentration in the hippocampus, cerebral cortex, or striatum. These results indicate that VPS13A has remarkable stability in neuronal cells. Understanding the distinct expression pattern of VPS13A can provide relevant information to unravel pathophysiological hallmarks of ChAc

Differential neuroprotective effects of 5'-deoxy-5'-methylthioadenosine

Background 5′-deoxy-5′-methylthioadenosine (MTA) is an endogenous compound produced through the metabolism of polyamines. The therapeutic potential of MTA has been assayed mainly in liver diseases and, more recently, in animal models of multiple sclerosis. The aim of this study was to determine the neuroprotective effect of this molecule in vitro and to assess whether MTA can cross the blood brain barrier (BBB) in order to also analyze its potential neuroprotective efficacy in vivo. Methods Neuroprotection was assessed in vitro using models of excitotoxicity in primary neurons, mixed astrocyte-neuron and primary oligodendrocyte cultures. The capacity of MTA to cross the BBB was measured in an artificial membrane assay and using an in vitro cell model. Finally, in vivo tests were performed in models of hypoxic brain damage, Parkinson's disease and epilepsy. Results MTA displays a wide array of neuroprotective activities against different insults in vitro. While the data from the two complementary approaches adopted indicate that MTA is likely to cross the BBB, the in vivo data showed that MTA may provide therapeutic benefits in specific circumstances. Whereas MTA reduced the neuronal cell death in pilocarpine-induced status epilepticus and the size of the lesion in global but not focal ischemic brain damage, it was ineffective in preserving dopaminergic neurons of the substantia nigra in the 1-methyl-4-phenyl-1,2,3,6-tetrahydro-pyr​idine(MPTP)-mice model. However, in this model of Parkinson's disease the combined administration of MTA and an A2A adenosine receptor antagonist did produce significant neuroprotection in this brain region. Conclusion MTA may potentially offer therapeutic neuroprotection

Differential neuroprotective effects of 5'-deoxy-5'-methylthioadenosine

Background 5′-deoxy-5′-methylthioadenosine (MTA) is an endogenous compound produced through the metabolism of polyamines. The therapeutic potential of MTA has been assayed mainly in liver diseases and, more recently, in animal models of multiple sclerosis. The aim of this study was to determine the neuroprotective effect of this molecule in vitro and to assess whether MTA can cross the blood brain barrier (BBB) in order to also analyze its potential neuroprotective efficacy in vivo. Methods Neuroprotection was assessed in vitro using models of excitotoxicity in primary neurons, mixed astrocyte-neuron and primary oligodendrocyte cultures. The capacity of MTA to cross the BBB was measured in an artificial membrane assay and using an in vitro cell model. Finally, in vivo tests were performed in models of hypoxic brain damage, Parkinson's disease and epilepsy. Results MTA displays a wide array of neuroprotective activities against different insults in vitro. While the data from the two complementary approaches adopted indicate that MTA is likely to cross the BBB, the in vivo data showed that MTA may provide therapeutic benefits in specific circumstances. Whereas MTA reduced the neuronal cell death in pilocarpine-induced status epilepticus and the size of the lesion in global but not focal ischemic brain damage, it was ineffective in preserving dopaminergic neurons of the substantia nigra in the 1-methyl-4-phenyl-1,2,3,6-tetrahydro-pyr​idine(MPTP)-mice model. However, in this model of Parkinson's disease the combined administration of MTA and an A2A adenosine receptor antagonist did produce significant neuroprotection in this brain region. Conclusion MTA may potentially offer therapeutic neuroprotection