Sleep in vitro : Erk pathway regulates sleep duration and sleep related genes

To date, for most biological and physiological phenomena, the scientific community has reach a consensus on their related function, except for sleep, which has an undetermined, albeit mystery, function. To further our understanding of sleep function(s), we first focused on the level of complexity at which sleep-like phenomenon can be observed. This lead to the development of an in vitro model. The second approach was to understand the molecular and cellular pathways regulating sleep and wakefulness, using both our in vitro and in vivo models. The third approach (ongoing) is to look across evolution when sleep or wakefulness appears. (1) To address the question as to whether sleep is a cellular property and how this is linked to the entire brain functioning, we developed a model of sleep in vitro by using dissociated primary cortical cultures. We aimed at simulating the major characteristics of sleep and wakefulness in vitro. We have shown that mature cortical cultures display a spontaneous electrical activity similar to sleep. When these cultures are stimulated by waking neurotransmitters, they show a tonic firing activity, similar to wakefulness, but return spontaneously to the "sleep-like" state 24h after stimulation. We have also shown that transcriptional, electrophysiological, and metabolic correlates of sleep and wakefulness can be reliably detected in dissociated cortical cultures. (2) To further understand at which molecular and cellular levels changes between sleep and wakefulness occur, we have used a pharmacological and systematic gene transcription approach in vitro and discovered a major role played by the Erk pathway. Indeed, pharmacological inhibition of this pathway in living animals decreased sleep by 2 hours per day and consolidated both sleep and wakefulness by reducing their fragmentation. (3) Finally, we tried to evaluate the presence of sleep in one of the most primitive species with a neural network. We set up Hydra as a model organism. We hypothesized that sleep as a cellular (neuronal) property may occur with the appearance of the most primitive nervous system. We were able to show that Hydra have periodic rest phases amounting to up to 5 hours per day. In conclusion, our work established an in vitro model to study sleep, discovered one of the major signaling pathways regulating vigilance states, and strongly suggests that sleep is a cellular property highly conserved at the molecular level during evolution. -- Jusqu'à ce jour, la communauté scientifique s'est mise d'accord sur la fonction d'une majorité des processus physiologiques, excepté pour le sommeil. En effet, la fonction du sommeil reste un mystère, et aucun consensus n'est atteint le concernant. Pour mieux comprendre la ou les fonctions du sommeil, (1) nous nous sommes d'abord concentré sur le niveau de complexité auquel un état ressemblant au sommeil peut être observé. Nous avons ainsi développé un modèle du sommeil in vitro, (2) nous avons disséqué les mécanismes moléculaires et cellulaires qui pourraient réguler le sommeil, (3) nous avons cherché à savoir si un état de sommeil peut être trouvé dans l'hydre, l'animal le plus primitif avec un système nerveux. (1) Pour répondre à la question de savoir à quel niveau de complexité apparaît un état de sommeil ou d'éveil, nous avons développé un modèle du sommeil, en utilisant des cellules dissociées de cortex. Nous avons essayé de reproduire les corrélats du sommeil et de l'éveil in vitro. Pour ce faire, nous avons développé des cultures qui montrent les signes électrophysiologiques du sommeil, puis quand stimulées chimiquement passent à un état proche de l'éveil et retournent dans un état de sommeil 24 heures après la stimulation. Notre modèle n'est pas parfait, mais nous avons montré que nous pouvions obtenir les corrélats électrophysiologiques, transcriptionnels et métaboliques du sommeil dans des cellules corticales dissociées. (2) Pour mieux comprendre ce qui se passe au niveau moléculaire et cellulaire durant les différents états de vigilance, nous avons utilisé ce modèle in vitro pour disséquer les différentes voies de signalisation moléculaire. Nous avons donc bloqué pharmacologiquement les voies majeures. Nous avons mis en évidence la voie Erkl/2 qui joue un rôle majeur dans la régulation du sommeil et dans la transcription des gènes qui corrèlent avec le cycle veille-sommeil. En effet, l'inhibition pharmacologique de cette voie chez la souris diminue de 2 heures la quantité du sommeil journalier et consolide l'éveil et le sommeil en diminuant leur fragmentation. (3) Finalement, nous avons cherché la présence du sommeil chez l'Hydre. Pour cela, nous avons étudié le comportement de l'Hydre pendant 24-48h et montrons que des périodes d'inactivité, semblable au sommeil, sont présentes dans cette espèce primitive. L'ensemble de ces travaux indique que le sommeil est une propriété cellulaire, présent chez tout animal avec un système nerveux et régulé par une voie de signalisation phylogénétiquement conservée.

Régulation transcriptionelle du développement de l'hypothalamus chez l'amphibien

Le noyau paraventriculaire (PVN) de l'hypothalamus régule une série de phénomènes physiologiques incluant l'équilibre énergétique et la pression artérielle. Nous avons identifié une cascade de facteurs de transcription qui contrôle le développement du PVN. SIM1 et OTP agissent en parallèle pour contrôler la différenciation d'au moins cinq types de neurones identifiables par la production d'OT, AVP, CRH, SS et TRH. Ces Facteurs de transcriptions contrôlent le développement des lignées CRH, AVP et OT en maintenant l'expression de Brn2 qui à son tour est nécessaire pour la différenciation terminale de ces neurones. L'analyse du transcriptome du PVN nous a permis d'identifier plusieurs gènes qui ont le potentiel de contrôler le développement du PVN. Nous voulons développer un paradigme de perte de fonction qui permettrait l'étude de ces gènes candidats sur une grande échelle. Le but de ce projet est de caractériser le PVN en développement de l'amphibien en vue de l'utilisation de ce modèle pour des études fonctionnelles. Nous avons cloné des fragments de cDNA de Sim1, OTP, Brn2, Sim2, CRH, Ot, AVP et TRH à partir de l'ARN total de Xenopus Laevis. Nous avons adapté notre technique d'hybridation in situ pour caractériser l'expression de ces gènes chez l'amphibien aux stades 33-39, 44, 51, 54, 60, et chez l'adulte. Résultats. Les Facteurs de transcription Sim1, OTP, et Brn2 commencent à être exprimés dans le PVN prospectif au stade 33. L'expression des marqueurs de différenciation terminale devient détectable entre les stades 37 et 39. De façon intéressante, le PVN occupe initialement un domaine de forme globulaire puis à partir du stade 44 s'allonge le long de l’axe dorso-ventral. Cet allongement se traduit par une organisation en colonnes des cellules du PVN que nous n'avons pas observée chez les rongeurs. Le développement du PVN est conservé chez l'amphibien dans la mesure où la relation entre l'expression des facteurs de transcription et des marqueurs de différenciation terminale est conservée. Il existe par ailleurs des différences entre la topographie des PVN des mammifères et de l'amphibien. L'organisation en colonnes de cellules pourrait correspondre à des mouvements de migration tangentielle. Nous sommes maintenant en mesure de tester la fonction des facteurs de transcription dans le PVN par l'approche d'invalidation par morpholinos.

Prevalencia de síntomas osteomusculares en miembros superiores y los probables factores de riesgo asociados en los estudiantes de musica en una institución universitaria de Bogotá,Colombia, 2013

Los músicos por su práctica instrumental tienen una alta demanda de desempeño físico, especialmente de los miembros superiores y están expuestos a varios factores de riesgo biomecánico que pueden resultar en problemas de salud. Objetivo: determinar la prevalencia de sintomatología osteomuscular de miembros superiores y los probables factores de riesgo asociados, en los estudiantes expuestos a la actividad musical durante el segundo semestre del año 2013 en una institución universitaria de Bogotá, Colombia. Método: se realizó un estudio descriptivo de corte trasversal en 134 estudiantes de todos los semestres de música en una institución universitaria. Se aplicó el Cuestionario nórdico estandarizado para análisis de síntomas músculo esqueléticos y una encuesta ad hoc que contemplaba aspectos sociodemográficos y antecedentes académicos, patológicos, factores de exposición y hábitos. Resultados: Las prevalencias generales encontradas en el estudio, son similares a las que refieren algunos estudios revisados que contemplan ciertas variables afines a las que se estudiaron. La prevalencia de síntomas osteomusculares cervico-braquial fue de 77.9%. La prevalencia de molestias en cuello fue mayor en las mujeres (64.3%) que en los hombres (37.4%) (OR=3.02, IC 95%=1.26, 7.18). La prevalencia de síntomas en manos/muñecas que le impidió hacer su trabajo en los últimos 12 meses fue mayor en los estudiantes que refirieron alguna enfermedad (29.4%) que en los que no la manifestaron (10.2%), (OR=3.69, IC 95%=1.34, 10.19). La prevalencia de molestias en cuello que les impidió hacer su trabajo en los últimos 12 meses fue mayor en los estudiantes que practicaron algún pasatiempo con sus brazos (10.4%) versus los que no lo practicaron, cuya frecuencia fue 0.0%. Los instrumentos musicales de mayor práctica fueron cuerda y percusión y se asociaron a prevalencia de síntomas osteomusculares cérvico-braquiales con una distribución por segmentos similar. Los tiempos de práctica semanales y la antigüedad en la práctica, conduce a síntomas cervico-braquiales. Conclusiones: Este estudio coincide con la distribución de las prevalencias encontradas en poblaciones de estudiantes de música revisadas, con respecto a la sintomatología, a los segmentos cervico-braquiales de mayor afectación, a la significancia del género femenino con respecto al masculino, al tipo de instrumentos y a los tiempos de práctica entre otros. Esto plantea la necesidad de educar a nuestros músicos en la detección temprana de síntomas desde su formación de pregrado o quizás mucho antes.

Maternal overnutrition programs changes in the expression of skeletal muscle genes that are associated with insulin resistance and defects of oxidative phosphorylation in adult male rat offspring.

Children of obese mothers have increased risk of metabolic syndrome as adults. Here we report the effects of a high-fat diet in the absence of maternal obesity at conception on skeletal muscle metabolic and transcriptional profiles of adult male offspring. Female Sprague Dawley rats were fed a diet rich in saturated fat and sucrose [high-fat diet (HFD): 23.5% total fat, 9.83% saturated fat, 20% sucrose wt:wt] or a normal control diet [(CD) 7% total fat, 0.5% saturated fat, 10% sucrose wt:wt] for the 3 wk prior to mating and throughout pregnancy and lactation. Maternal weights were not different at conception; however, HFD-fed dams were 22% heavier than controls during pregnancy. On a normal diet, the male offspring of HFD-fed dams were not heavier than controls but demonstrated features of insulin resistance, including elevated plasma insulin concentration [40.1 ± 2.5 (CD) vs 56.2 ± 6.1 (HFD) mU/L; P = 0.023]. Next-generation mRNA sequencing was used to identify differentially expressed genes in the offspring soleus muscle, and gene set enrichment analysis (GSEA) was used to detect coordinated changes that are characteristic of a biological function. GSEA identified 15 upregulated pathways, including cytokine signaling (P < 0.005), starch and sucrose metabolism (P < 0.017), inflammatory response (P < 0.024), and cytokine-cytokine receptor interaction (P < 0.037). A further 8 pathways were downregulated, including oxidative phosphorylation (P < 0.004), mitochondrial matrix (P < 0.006), and electron transport/uncoupling (P < 0.022). Phosphorylation of the insulin signaling protein kinase B was reduced [2.86 ± 0.63 (CD) vs 1.02 ± 0.27 (HFD); P = 0.027] and mitochondrial complexes I, II, and V protein were downregulated by 50-68% (P < 0.005). On a normal diet, the male offspring of HFD-fed dams did not become obese adults but developed insulin resistance, with transcriptional evidence of muscle cytokine activation, inflammation, and mitochondrial dysfunction. These data indicate that maternal overnutrition, even in the absence of prepregnancy obesity, can promote metabolic dysregulation and predispose offspring to type 2 diabetes.

Observable essential fatty acid deficiency markers and autism spectrum disorder

Autism Spectrum Disorder (ASD) has been associated with essential fatty acid (EFA) deficiencies, with some researchers theorising that dysregulation of phospholipid metabolism may form part of the biological basis for ASD. This pilot study compared observable signs of fatty acid status of 19 children with an ASD diagnosis to 23 of their typically developing siblings. A pregnancy, birth and breastfeeding history was also obtained from their parents, which included a measure of infant intake of fatty acid rich colostrum immediately post-partum. When considered within their family group, those infants not breastfed (with colostrum) within the first hour of life and who had a history of fatty acid deficiency symptoms were more likely to have an ASD diagnosis. Other variables such as formula use, duration of breastfeeding, gestational age and Apgar scores were not associated with group membership. The results of this study are consistent with previous research showing a relationship between fatty acid metabolism, breastfeeding and ASD such that early infant feeding practices and the influence this has on the fatty acid metabolism of the child may be a risk factor for ASD.

Differences in dietary pattern between obese and eutrophic children

Background: Excessive consumption of energy is a decisive factor of obesity, but a simple quantitative assessment of consumption between obese and eutrophic individuals not always explains the problem, raising questions about the importance of the qualitative aspects of food. Therefore, the purpose of this study was to evaluate the differences in nutrient composition and meal patterns between eutrophic and obese schoolchildren. Methods. The diet of 83 children (42 obese and 41 eutrophic), aged between 7 and 11 years of age, was assessed by two non-consecutive dietary recalls. After the software analysis of macro and micronutrients composition, the different types and amount of legumes, fruits and vegetables were analyzed to verify the dietary patterns. Results: No differences were verified in energy consumption between the groups (eutrophic = 1934.2 672.7 kcal, obese = 1835.8 621.2 kcal). In general, children showed consumption within the recommended ranges of carbohydrates, lipids and proteins. The average consumption of fiber was higher in the eutrophic group (20.7 g) when compared to the obese group (14.8 g). The dietary fiber was strongly correlated with the number of servings of beans (r = 0.77), when compared to fruits (r = 0.44) and leafy vegetables (r = 0.13). It was also observed that the higher the consumption of fiber and beans, the lower the proportion of dietary fat (r = -0.22) in the diet. Generally, there was a low consumption of fiber (20.7 g = eutrophic group/14.8 g = obese group), beans (1.1 portions in the eutrophic and obese groups), fruits (0.7 portions eutrophic group and 0.6 obese group) and vegetables (1.3 eutrophic group and 1.1 obese group). Conclusions: It is concluded that the obesity was more related to a dietary pattern of low intake of dietary fiber than excessive energy consumption and macronutrients imbalance. © 2011 de Oliveira et al; licensee BioMed Central Ltd.

Meat quality of young Nellore bulls with low and high residual feed intake

Fifty-nine Nellore bulls from low and high residual feed intake (RFI) levels were studied with the objective of evaluating meat quality traits. Animals were slaughtered when ultrasound-measured backfat thickness reached 4. mm, and samples of Longissimus were collected. A mixed model including RFI as fixed effect and herd and diet as random effects was used, and least square means were compared by t-test. More efficient animals consumed 0.730. kg dry matter/day less than less efficient animals, with similar performance. No significant differences in carcass weight, prime meat cuts proportion, chemical composition, pH, sarcomere length, or color were observed between RFI groups. Shear force, myofibrillar fragmentation index and soluble collagen content were influenced by RFI, with a higher shear force and soluble collagen content and a lower fragmentation index in low RFI animals. Feedlot-finished low RFI young Nellore bulls more efficiently convert feed into meat, presenting carcasses within quality standards. © 2012 Elsevier Ltd.

Cardiorespiratory fitness, but not central obesity or C-reactive protein, is related to liver function in obese children

Nonalcoholic fatty liver disease (NAFLD) is one of the most frequent complications associated with excess adiposity. Its pathogenesis is complex and there are multiple factors that may contribute to it. AIM: To analyze whether cardiorespiratory ftness (CRF), waist circumference (WC), and C-reactive protein (CRP) are associated with alanine aminotransferase (ALT) in children with obesity. METHODS: 79 overweight/obese children of both genders, 11-13 year-olds, with abnormal serum ALT from Porto public schools comprised the sample. Measurements included CRF (20-m Shuttle Run Test), WC (NHANES protocol), CRP and ALT (Cholestech LDX analyzer). Logistic regression adjusted for gender, maturation, and weight with ALT levels as dependent variable (risk vs. non risk), and WC (risk vs. non risk), CRP (risk vs. non risk), and CRF (fit vs. unfit) as independent variables. Level of significance was set at 95%. RESULTS: Logistic regression showed that obese fit children were less likely to have abnormal ALT values (OR=.031) CONCLUSION: In obese children, higher cardiovascular fitness appears to reduce the chance of decreased liver function. © 2013 Human Kinetics, Inc.

Effects of corn replacement by sorghum in broiler diets on performance and intestinal mucosa integrity

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Maternal protein restriction during pregnancy affects gene expression and immunolocalization of intestinal nutrient transporters in rats

Intrauterine dietary restriction may cause changes in the functioning of offspring organs and systems later in life, an effect known as fetal programming. The present study evaluated mRNA abundance and immunolocalization of nutrient transporters as well as enterocytes proliferation in the proximal, median and distal segments of small intestine of rats born to protein-restricted dams. Pregnant rats were fed hypoproteic (6% protein) or control (17% protein) diets, and offspring rats were evaluated at 3 and 16 weeks of age. The presence of SGLT1 (sodium-glucose co-transporter 1), GLUT2 (glucose transporter 2), PEPT1 (peptide transporter 1) and the intestinal proliferation were evaluated by immunohistochemical techniques and the abundance of specific mRNA for SGLT1, GLUT2 and PEPT1 was assessed by the real-time PCR technique. Rats born to protein-restricted dams showed higher cell proliferation in all intestinal segments and higher gene expression of SGLT1 and PEPT1 in the duodenum. Moreover, in adult animals born to protein-restricted dams the immunoreactivity of SGLT1, GLUT2 and PEPT1in the duodenum was more intense than in control rats. Taken together, the results indicate that changes in the small intestine observed in adulthood can be programmed during the gestation. In addition, they show that this response is caused by both up-regulation in transporter gene expression, a specific adaptation mechanism, and intestinal proliferation, an unspecific adaptation mechanism.

Caracterização sistêmica de alterações em redes fisiológicas de plantas C3 e C4 submetidas à deficiência hídrica

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Temporal analysis of prostaglandin F2 alpha receptor, caspase 3, and cyclooxygenase 2 messenger RNA expression and prostaglandin F2 alpha receptor and cyclooxygenase 2 protein expression in endometrial tissue from multiparous Nelore (Bos taurus indicus) cows treated with cloprostenol sodium during puerperium

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estratégias eletrolíticas: controle do estresse térmico em ovinos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Ethnic/racial diversity, maternal stress, lactation and very low birthweight infants.

OBJECTIVE: (1) To compare maternal characteristics and psychological stress profile among African-American, Caucasian and Hispanic mothers who delivered very low birthweight infants. (2) To investigate associations between psychosocial factors, frequency of milk expression, skin-to-skin holding (STS), and lactation performance, defined as maternal drive to express milk and milk volume. STUDY DESIGN: Self-reported psychological questionnaires were given every 2 weeks after delivery over 10 weeks. Milk expression frequency, STS, and socioeconomic variables were collected. RESULT: Infant birthweight, education, and milk expression frequency differed between groups. Trait anxiety, depression and parental stress in a neonatal intensive care unit (PSS:NICU) were similar. African-American and Caucasian mothers reported the lowest scores in state anxiety and social desirability, respectively. Maternal drive to express milk, measured by maintenance of milk expression, correlated negatively with parental role alteration (subset of PSS:NICU) and positively with infant birthweight and STS. Milk volume correlated negatively with depression and positively with milk expression frequency and STS. CONCLUSION: Differences between groups were observed for certain psychosocial factors. The response bias to self-reported questionnaires between groups may not provide an accurate profile of maternal psychosocial profile. With different factors correlating with maintenance of milk expression and milk volume, lactation performance can be best enhanced with a multi-faceted intervention program, incorporating parental involvement in infant care, close awareness and management of maternal mental health, and encouragement for frequent milk expression and STS.

Functional taxonomy of bacterial hyperstructures.

The levels of organization that exist in bacteria extend from macromolecules to populations. Evidence that there is also a level of organization intermediate between the macromolecule and the bacterial cell is accumulating. This is the level of hyperstructures. Here, we review a variety of spatially extended structures, complexes, and assemblies that might be termed hyperstructures. These include ribosomal or "nucleolar" hyperstructures; transertion hyperstructures; putative phosphotransferase system and glycolytic hyperstructures; chemosignaling and flagellar hyperstructures; DNA repair hyperstructures; cytoskeletal hyperstructures based on EF-Tu, FtsZ, and MreB; and cell cycle hyperstructures responsible for DNA replication, sequestration of newly replicated origins, segregation, compaction, and division. We propose principles for classifying these hyperstructures and finally illustrate how thinking in terms of hyperstructures may lead to a different vision of the bacterial cell.