Meta-analysis and imputation refines the association of 15q25 with smoking quantity.

Smoking is a leading global cause of disease and mortality. We established the Oxford-GlaxoSmithKline study (Ox-GSK) to perform a genome-wide meta-analysis of SNP association with smoking-related behavioral traits. Our final data set included 41,150 individuals drawn from 20 disease, population and control cohorts. Our analysis confirmed an effect on smoking quantity at a locus on 15q25 (P = 9.45 x 10(-19)) that includes CHRNA5, CHRNA3 and CHRNB4, three genes encoding neuronal nicotinic acetylcholine receptor subunits. We used data from the 1000 Genomes project to investigate the region using imputation, which allowed for analysis of virtually all common SNPs in the region and offered a fivefold increase in marker density over HapMap2 (ref. 2) as an imputation reference panel. Our fine-mapping approach identified a SNP showing the highest significance, rs55853698, located within the promoter region of CHRNA5. Conditional analysis also identified a secondary locus (rs6495308) in CHRNA3.

Six new loci associated with body mass index highlight a neuronal influence on body weight regulation.

Common variants at only two loci, FTO and MC4R, have been reproducibly associated with body mass index (BMI) in humans. To identify additional loci, we conducted meta-analysis of 15 genome-wide association studies for BMI (n > 32,000) and followed up top signals in 14 additional cohorts (n > 59,000). We strongly confirm FTO and MC4R and identify six additional loci (P < 5 x 10(-8)): TMEM18, KCTD15, GNPDA2, SH2B1, MTCH2 and NEGR1 (where a 45-kb deletion polymorphism is a candidate causal variant). Several of the likely causal genes are highly expressed or known to act in the central nervous system (CNS), emphasizing, as in rare monogenic forms of obesity, the role of the CNS in predisposition to obesity.

Identification of the pathological alterations underlying respiratory failure in myotonic dystrophy transgenic mice

AbstractMyotonic dystrophy type 1 (DM1), also known as Steinert's disease, is an inherited autosomal dominant disease. DM1 is characterized by myotonia, muscular weakness and atrophy, but it has a multisystemic phenotype. The genetic basis of the disease is the abnormal expansion of CTG repeats in the 3' untranslated region of the DM protein kinase (DMPK) gene on chromosome 19. The size of the expansion correlates to the severity of the disease and the age of onset.Respiratory problems have long been recognized to be a major feature of the disease and are the main factor contributing to mortality ; however the mechanisms are only partly known. The aim of our study is to investigate whether respiratory failure results only from the involvement of the dystrophic process at the level of the respiratory muscles or comes also from abnormalities in the neuronal network that generates and controls the respiratory rhythm. The generation of valid transgenic mice displaying the human DM1 phenotype by the group of Dr. Gourdon provided us a useful tool to analyze the brain stem respiratory neurons, spinal phrenic motoneurons and phrenic nerves. We examined therefore these structures in transgenic mice carrying 350-500 CTGs and displaying a mild form of the disease (DM1 mice). The morphological and morphometric analysis of diaphragm muscle sections revealed a denervation of the end-plates (EPs), characterized by a decrease in size and shape complexity of EPs and a reduction in the density of acetylcholine receptors (AChRs). Also a strong and significant reduction in the number of phrenic unmyelinated fibers was detected, but not in the myelinated fibers. In addition, no pathological changes were detected in the cervical motoneurons and medullary respiratory centers (Panaite et al., 2008). These results suggest that the breathing rhythm is probably not affected in mice expressing a mild form of DM1, but rather the transmission of action potentials at the level of diaphragm NMJs is deficient.Because size of the mutation increases over generations, new transgenic mice were obtained from the mice with 350-500 CTGs, resulting from a large increase of CTG repeat in successive generations, these mice carry more than 1300 CTGs (DMSXL) and display a severe DM1 phenotype (Gomes-Pereira et al., 2007). Before we study the mechanism underlying the respiratory failure in DMSXL mice, we analyzed the peripheral nervous system (PNS) in these mice by electrophysiological, histological and morphometric methods. Our results provide strong evidence that DMSXL mice have motor neuropathy (Panaite et al., 2010, submitted). Therefore the DMSXL mice expressing severe DM1 features represent for us a good tool to investigate, in the future, the physiological, structural and molecular alterations underlying respiratory failure in DM1. Understanding the mechanism of respiratory deficiency will help to better target the therapy of these problems in DM1 patients. In addition our results may, in the future, orientate pharmaceutical and clinical research towards possible development of therapy against respiratory deficits associated with the DM1.RésuméLa dystrophic myotonique type 1 (DM1), aussi dénommée maladie de Steinert, est une maladie héréditaire autosomique dominante. Elle est caractérisée par une myotonie, une faiblesse musculaire avec atrophie et se manifeste aussi par un phénotype multisystémique. La base génétique de la maladie est une expansion anormale de répétitions CTG dans une région non traduite en 3' du gène de la DM protéine kinase (DMPK) sur le chromosome 19. La taille de l'expansion est corrélée avec la sévérité et l'âge d'apparition de DM1.Bien que les problèmes respiratoires soient reconnus depuis longtemps comme une complication de la maladie et soient le principal facteur contribuant à la mortalité, les mécanismes en sont partiellement connus. Le but de notre étude est d'examiner si l'insuffisance respiratoire de la DM1 est dû au processus dystrophique au niveau des muscles respiratoires ou si elle est entraînée aussi par des anomalies dans le réseau neuronal qui génère et contrôle le rythme respiratoire. La production par le groupe du Dr. Gourdon de souris transgéniques de DM1, manifestant le phénotype de DM1 humaine, nous a fourni un outil pour analyser les nerfs phréniques, les neurones des centres respiratoires du tronc cérébral et les motoneurones phréniques. Par conséquence, nous avons examiné ces structures chez des souris transgéniques portant 350-500 CTG et affichant une forme légère de la maladie (souris DM1). L'analyse morphologique et morphométrique des sections du diaphragme a révélé une dénervation des plaques motrices et une diminution de la taille et de la complexité de la membrane postsynaptîque, ainsi qu'une réduction de la densité des récepteurs à l'acétylcholine. Nous avons aussi détecté une réduction significative du nombre de fibres nerveuses non myélinisées mais pas des fibres myélinisées. Par ailleurs, aucun changement pathologique n'a été détecté pour les neurones moteurs médullaires cervicaux et centres respiratoires du tronc cérébral (Panaite et al., 2008). Ces résultats suggèrent que le iythme respiratoire n'est probablement pas affecté chez les souris manifestant une forme légère du DM1, mais plutôt que la transmission des potentiels d'action au niveau des plaques motrices du diaphragme est déficiente.Comme la taille du mutation augmente au fil des générations, de nouvelles souris transgéniques ont été générés par le groupe Gourdon; ces souris ont plus de 1300 CTG (DMSXL) et manifestent un phénotype sévère du DM1 (Gomes-Pereira et al., 2007). Avant d'étudier le mécanisme sous-jacent de l'insuffisance respiratoire chez les souris DMSXL, nous avons analysé le système nerveux périphérique chez ces souris par des méthodes électrophysiologiques, histologiques et morphométriques. Nos résultats fournissent des preuves solides que les souris DMSXL manifestent une neuropathie motrice (Panaite et al., 2010, soumis). Par conséquent, les souris DMSXL représentent pour nous un bon outil pour étudier, à l'avenir, les modifications physiologiques, morphologiques et moléculaires qui sous-tendent l'insuffisance respiratoire du DM1. La connaissance du mécanisme de déficience respiratoire en DM1 aidera à mieux cibler le traitement de ces problèmes aux patients. De plus, nos résultats pourront, à l'avenir, orienter la recherche pharmaceutique et clinique vers le développement de thérapie contre le déficit respiratoire associé à DM1.

Triodothyronine enhancement of neuronal differentiation in aggregating fetal rat brain cells cultured in a chemically defined medium.

Triiodothyronine (30 nM) added to serum-free cultures of mechanically dissociated re-aggregating fetal (15-16 days gestation) rat brain cells greatly increased the enzymatic activity of choline acetyltransferase and acetylcholinesterase throughout the entire culture period (33 days), and markedly accelerated the developmental rise of glutamic acid decarboxylase specific activity. The enhancement of choline acetyltransferase and acetylcholinesterase specific activities in the presence of triiodothyronine was even more pronouned in cultures of telencephalic cells. If triiodothyronine treatment was restricted to the first 17 culture days, the level of choline acetyltransferase specific activity at day 33 was 84% of that in chronically treated cultures and 270% of that in cultures receiving triiodothyronine between days 17 and 33, indicating that relatively undifferentiated cells were more responsive to the hormone. Triiodothyronine had no apparent effect on the incorporation of [3H]thymidine at day 5 or on the total DNA content of cultures, suggesting that cellular differentiation, rather than proliferation was affected by the hormone. Our findings in vitro are in good agreement with many observations in vivo, suggesting that rotation-mediated aggregating cell cultures of fetal rat brain provide a useful model to study thyroid hormone action in the developing brain.

Official Positions for FRAX® Bone Mineral Density and FRAX® simplification from Joint Official Positions Development Conference of the International Society for Clinical Densitometry and International Osteoporosis Foundation on FRAX®.

Tools to predict fracture risk are useful for selecting patients for pharmacological therapy in order to reduce fracture risk and redirect limited healthcare resources to those who are most likely to benefit. FRAX® is a World Health Organization fracture risk assessment algorithm for estimating the 10-year probability of hip fracture and major osteoporotic fracture. Effective application of FRAX® in clinical practice requires a thorough understanding of its limitations as well as its utility. For some patients, FRAX® may underestimate or overestimate fracture risk. In order to address some of the common issues encountered with the use of FRAX® for individual patients, the International Society for Clinical Densitometry (ISCD) and International Osteoporosis Foundation (IOF) assigned task forces to review the medical evidence and make recommendations for optimal use of FRAX® in clinical practice. Among the issues addressed were the use of bone mineral density (BMD) measurements at skeletal sites other than the femoral neck, the use of technologies other than dual-energy X-ray absorptiometry, the use of FRAX® without BMD input, the use of FRAX® to monitor treatment, and the addition of the rate of bone loss as a clinical risk factor for FRAX®. The evidence and recommendations were presented to a panel of experts at the Joint ISCD-IOF FRAX® Position Development Conference, resulting in the development of Joint ISCD-IOF Official Positions addressing FRAX®-related issues.

Neuroligin-1 links neuronal activity to sleep-wake regulation.

Maintaining wakefulness is associated with a progressive increase in the need for sleep. This phenomenon has been linked to changes in synaptic function. The synaptic adhesion molecule Neuroligin-1 (NLG1) controls the activity and synaptic localization of N-methyl-d-aspartate receptors, which activity is impaired by prolonged wakefulness. We here highlight that this pathway may underlie both the adverse effects of sleep loss on cognition and the subsequent changes in cortical synchrony. We found that the expression of specific Nlg1 transcript variants is changed by sleep deprivation in three mouse strains. These observations were associated with strain-specific changes in synaptic NLG1 protein content. Importantly, we showed that Nlg1 knockout mice are not able to sustain wakefulness and spend more time in nonrapid eye movement sleep than wild-type mice. These changes occurred with modifications in waking quality as exemplified by low theta/alpha activity during wakefulness and poor preference for social novelty, as well as altered delta synchrony during sleep. Finally, we identified a transcriptional pathway that could underlie the sleep/wake-dependent changes in Nlg1 expression and that involves clock transcription factors. We thus suggest that NLG1 is an element that contributes to the coupling of neuronal activity to sleep/wake regulation.

Effect of water velocity on the uptake of polychlorinatedd biphenyls (PCBs) by silicone rubber (SR) and low-density polyethylene (LDPE) passive samplers: an assessment of the efficiency of performance reference compounds (PRCs) in river-like flow conditions

One aim of this study is to determine the impact of water velocity on the uptake of indicator polychlorinated biphenyls (iPCBs) by silicone rubber (SR) and low-density polyethylene (LDPE) passive samplers. A second aim is to assess the efficiency of performance reference compounds (PRCs) to correct for the impact of water velocity. SR and LDPE samplers were spiked with 11 or 12 PRCs and exposed for 6 weeks to four different velocities (in the range of 1.6 to 37.7 cm s−1) in river-like flow conditions using a channel system supplied with river water. A relationship between velocity and the uptakewas found for each iPCB and enables to determine expected changes in the uptake due to velocity variations. For both samplers, velocity increases from 2 to 10 cm s−1, 30 cm s−1 (interpolated data) and 100 cm s−1 (extrapolated data) lead to increases of the uptake which do not exceed a factor of 2, 3 and 4.5, respectively. Results also showed that the influence of velocity decreased with increasing the octanol-water coefficient partition (log Kow) of iPCBs when SR is used whereas the opposite effect was observed for LDPE. Time-weighted average (TWA) concentrations of iPCBs in water were calculated from iPCB uptake and PRC release. These calculations were performed using either a single PRC or all the PRCs. The efficiency of PRCs to correct the impact of velocity was assessed by comparing the TWA concentrations obtained at the four tested velocities. For SR, a good agreement was found among the four TWA concentrations with both methods (average RSD b 10%). Also for LDPE, PRCs offered a good correction of the impact of water velocity (average RSD of about 10 to 20%). These results contribute to the process of acceptance of passive sampling in routine regulatory monitoring programs.

Lack of GDAP1 induces neuronal calcium and mitochondrial defects in a knockout mouse model of charcot-marie-tooth neuropathy.

Mutations in GDAP1, which encodes protein located in the mitochondrial outer membrane, cause axonal recessive (AR-CMT2), axonal dominant (CMT2K) and demyelinating recessive (CMT4A) forms of Charcot-Marie-Tooth (CMT) neuropathy. Loss of function recessive mutations in GDAP1 are associated with decreased mitochondrial fission activity, while dominant mutations result in impairment of mitochondrial fusion with increased production of reactive oxygen species and susceptibility to apoptotic stimuli. GDAP1 silencing in vitro reduces Ca2+ inflow through store-operated Ca2+ entry (SOCE) upon mobilization of endoplasmic reticulum (ER) Ca2+, likely in association with an abnormal distribution of the mitochondrial network. To investigate the functional consequences of lack of GDAP1 in vivo, we generated a Gdap1 knockout mouse. The affected animals presented abnormal motor behavior starting at the age of 3 months. Electrophysiological and biochemical studies confirmed the axonal nature of the neuropathy whereas histopathological studies over time showed progressive loss of motor neurons (MNs) in the anterior horn of the spinal cord and defects in neuromuscular junctions. Analyses of cultured embryonic MNs and adult dorsal root ganglia neurons from affected animals demonstrated large and defective mitochondria, changes in the ER cisternae, reduced acetylation of cytoskeletal α-tubulin and increased autophagy vesicles. Importantly, MNs showed reduced cytosolic calcium and SOCE response. The development and characterization of the GDAP1 neuropathy mice model thus revealed that some of the pathophysiological changes present in axonal recessive form of the GDAP1-related CMT might be the consequence of changes in the mitochondrial network biology and mitochondria-endoplasmic reticulum interaction leading to abnormalities in calcium homeostasis.

Proximity of excitatory synapses and astroglial gap junctions in layer IV of the mouse barrel cortex.

Neurons and astrocytes, the two major cell populations in the adult brain, are characterized by their own mode of intercellular communication--the synapses and the gap junctions (GJ), respectively. In addition, there is increasing evidence for dynamic and metabolic neuroglial interactions resulting in the modulation of synaptic transmission at the so-called "tripartite synapse". Based on this, we have investigated at the ultrastructural level how excitatory synapses (ES) and astroglial GJ are spatially distributed in layer IV of the barrel cortex of the adult mouse. We used specific antibodies for connexin (Cx) 30 and 43 to identify astroglial GJ, these two proteins are known to be present in the majority of astroglial GJ in the cerebral cortex. In electron-microscopic images, we measured the distance between two ES, between two GJ and between a GJ and its nearest ES. We found a ratio of two GJ per three ES in the hollow and septal areas. Taking into account the size of an astrocyte domain, the high density of GJ suggests the occurrence of reflexive type, i.e. GJ between processes of the same astrocyte. Interestingly, the distance between an ES and an astroglial GJ was found to be significantly lower than that between either two synapses or between two GJ. These observations indicate that the two modes of cell-to-cell communication are not randomly distributed in layer IV of the barrel cortex. Consequently, this feature may provide the morphological support for the recently reported functional interactions between neuronal circuits and astroglial networks.

Review of quantitative phase-digital holographic microscopy: promising novel imaging technique to resolve neuronal network activity and identify cellular biomarkers of psychiatric disorders.

Quantitative phase microscopy (QPM) has recently emerged as a new powerful quantitative imaging technique well suited to noninvasively explore a transparent specimen with a nanometric axial sensitivity. In this review, we expose the recent developments of quantitative phase-digital holographic microscopy (QP-DHM). Quantitative phase-digital holographic microscopy (QP-DHM) represents an important and efficient quantitative phase method to explore cell structure and dynamics. In a second part, the most relevant QPM applications in the field of cell biology are summarized. A particular emphasis is placed on the original biological information, which can be derived from the quantitative phase signal. In a third part, recent applications obtained, with QP-DHM in the field of cellular neuroscience, namely the possibility to optically resolve neuronal network activity and spine dynamics, are presented. Furthermore, potential applications of QPM related to psychiatry through the identification of new and original cell biomarkers that, when combined with a range of other biomarkers, could significantly contribute to the determination of high risk developmental trajectories for psychiatric disorders, are discussed.

3-D density structure and geological evolution of Stromboli volcano (Aeolian Islands, Italy) inferred from land-based and sea-surface gravity data

We present the first density model of Stromboli volcano (Aeolian Islands, Italy) obtained by simultaneously inverting land-based (543) and sea-surface (327) relative gravity data. Modern positioning technology, a 1 x 1 m digital elevation model, and a 15 x 15 m bathymetric model made it possible to obtain a detailed 3-D density model through an iteratively reweighted smoothness-constrained least-squares inversion that explained the land-based gravity data to 0.09 mGal and the sea-surface data to 5 mGal. Our inverse formulation avoids introducing any assumptions about density magnitudes. At 125 m depth from the land surface, the inferred mean density of the island is 2380 kg m(-3), with corresponding 2.5 and 97.5 percentiles of 2200 and 2530 kg m-3. This density range covers the rock densities of new and previously published samples of Paleostromboli I, Vancori, Neostromboli and San Bartolo lava flows. High-density anomalies in the central and southern part of the island can be related to two main degassing faults crossing the island (N41 and NM) that are interpreted as preferential regions of dyke intrusions. In addition, two low-density anomalies are found in the northeastern part and in the summit area of the island. These anomalies seem to be geographically related with past paroxysmal explosive phreato-magmatic events that have played important roles in the evolution of Stromboli Island by forming the Scari caldera and the Neostromboli crater, respectively. (C) 2014 Elsevier B.V. All rights reserved.

Targeted inactivation of hypocretin receptors in dopaminergic and noradrenergic systems alters brain activity in wakefulness

Since the discovery of hypocretins/orexins (Hcrt/Ox) in 1998, several narcoleptic mouse models, such as Hcrt-KO, Hcrtrl-KO, Hcrtr2-KO and double receptors KO mice, and orexin-ataxin transgenic mice were generated. The available Hcrt mouse models do not allow the dissection of the specific role of Hcrt in each target region. Dr. Anne Vassalli generated loxP-flanked alleles for each Hcrt receptor, which are manipulated by Cre recombinase to generate mouse lines with disrupted Hcrtrl or Hcrtr2 (or both) in cell type-specific manner. The role of noradrenaline (NA) and dopamine (OA) in ttie regulation of vigilance states is well documented. The purpose of this thesis is to explore the role of the Hcrt input into these two monoaminergic systems. Chronic loss of Hcrtrl in NA neurons consolidated paradoxical sleep (PS), and altered wakefulness brain activity in baseline, during the sleep deprivation (SD), and when mice were challenged by a novel environment, or exposed to nest-building material. The analysis of alterations in the sleep EEG delta power showed a consistent correlation with the changes in the preceding waking quality in these mice. Targeted inactivation of Hcrt input into DA neurons showed that Hcrtr2 inactivation present the strongest phenotype. The loss of Hcrtr2 in DA neurons caused modified brain activities in spontaneous wakefulness, during SD, and in novel environmental conditions. In addition to alteration of wakefulness quality and quantity, conditional inactivation of Hcrtr2 in DA neurons caused an increased in time spent in PS in baseline and a delayed and less complete PS recovery after SD. In the first 30 min of sleep recovery, single (i.e. for Hcrtrl or Hcrtr2) conditional knockout receptor mice had opposite changes in delta activity, including an increased power density in the fast delta range with specific inactivation of Hcrtr2, but a decreased power density in the same range with specific inactivation of Hcrtrl in DA cells. These studies demonstrate a complex impact of Hcrt receptors signaling in both NA and DA system, not only on quantity and quality of wakefulness, but also on PS amount regulation as well as on SWS delta power expression. -- Depuis la découverte des hypocrétines/orexines (Hcrt/Ox) en 1998, plusieurs modèles de souris, narcoleptiques telles que Hcrt-KO, Hcrtr2-KO et récepteurs doubles KO et les souris transgéniques orexine-ataxine ont été générés. Les modèles de souris Hcrt disponibles ne permettaient pas la dissection du rôle spécifique de l'Hcrt dans chaque noyau neuronal cible. Notre laboratoire a généré des allèles loxP pour chacun des 2 gènes codant pour les récepteurs Hcrtr, qui sont manipulés par recombinase Cre pour générer des lignées de souris avec Hcrtrl inactivé, ou Hcrtr2 inactivé, (ou les deux), spécifiquement dans un type cellulaire particulier. Le rôle de la noradrénaline (NA) et la dopamine (DA) dans la régulation des états de vigilance est bien documentée. Le but de cette thèse est d'étudier le rôle de l'afférence Hcrt dans ces deux systèmes monoaminergiques au niveau de l'activité cérébrale telle qu'elle apparaît dans l'électroencéphalogramme (EEG). Mon travail montre que la perte chronique de Hcrtrl dans les neurones NA consolide le sommeil paradoxal (PS), et l'activité cérébrale de l'éveil est modifiée en condition spontanée, au cours d'une experience de privation de sommeil (SD), et lorsque les souris sont présentées à un nouvel environnement, ou exposées à des matériaux de construction du nid. Ces modifications de l'éveil sont corrélées à des modifications de puissance de l'activité delta du sommeil lent qui le suit. L'inactivation ciblée des Hcrtrs dans les neurones DA a montré que l'inactivation Hcrtr2 conduit au phénotype le plus marqué. La perte de Hcrtr2 dans les neurones DA mène à des modification d'activité cérébrale en éveil spontané, pendant SD, ainsi que dans des conditions environnementales nouvelles. En plus de l'altération de la qualité de l'éveil et de la quantité, l'inactivation conditionnelle de Hcrtr2 dans les neurones DA a provoqué une augmentation du temps passé en sommeil paradoxal (PS) en condition de base, et une reprise retardée et moins complète du PS après SD. Dans les 30 premières minutes de la récupération de sommeil, les modèles inactivés pour un seul des récepteurs (ie pour Hcrtrl ou Hcrtr2 seulement) montrent des changements opposés en activité delta, en particulier une densité de puissance accrue dans le delta rapide avec l'inactivation spécifique de Hcrtr2, mais une densité de puissance diminuée dans cette même gamme chez les souris inactivées spécifiquement en Hcrtrl dans les neurones DA. Ces études démontrent un impact complexe de l'inactivation de la neurotransmission au niveau des récepteurs d'Hcrt dans les deux compartiments NA et DA, non seulement sur la quantité et la qualité de l'éveil, mais aussi sur la régulation de quantité de sommeil paradoxal, ainsi que sur l'expression de la puissance delta pendant le sommeil lent.