Comparative winter thermoregulation and body temperature in three sympatric Apodemus species (A-alpicola, A-flavicollis, and A-sylvaticus)

When living in sympatry with Apodemus sylvaticus and A. flavicollis, A. alpicola dominates numerically at higher altitudes. A more efficient winter thermal isolation or a higher winter thermogenic capacity procuring a physiological advantage could explain at least part of this domination. We therefore measured body temperature (Tb), oxygen consumption (VO2), wet minimal thermal conductance (C) and non shivering thermogenesis (NST) at different ambient temperatures (Ta) on winter acclimated mice of the three species, and this for the first time in A. alpicola. NST was high and C low in the three species. No significant difference could be noticed either in Tb between 5 and -10 degrees C, in VO2 measurements at a Ta of -10 degrees C or in C. The NST measurements represent, respectively, 135.2% for A. sylvaticus, 142.8% for A. flavicollis and 140.5% for A. alpicola of the expected values, the values for A. sylvaticus being significantly lower than for the other two species. The basal metabolic rates (BMR) represent 169.4% for A. sylvaticus, 161.6% for A. flavicollis and 138.3% for A. alpicola of the expected values. Having removed the effect of body weight, the BMR value was significantly lower in A. alpicola than in A. flavicollis, but no difference could be noticed between A. sylvaticus and the other two species. In conclusion, the three species of mice have very similar acclimated thermoregulatory characteristics, well adapted to cold ambient conditions. One discriminating and advantageous factor could be the lower basal metabolic rate measured in A. alpicola compared to the other two species.

Effects of supplementation with essential amino acids on intrahepatic lipid concentrations during fructose overfeeding in humans.

BACKGROUND: A high dietary protein intake has been shown to blunt the deposition of intrahepatic lipids in high-fat- and high-carbohydrate-fed rodents and humans. OBJECTIVE: The aim of this study was to evaluate the effect of essential amino acid supplementation on the increase in hepatic fat content induced by a high-fructose diet in healthy subjects. DESIGN: Nine healthy male volunteers were studied on 3 occasions in a randomized, crossover design after 6 d of dietary intervention. Dietary conditions consisted of a weight-maintenance balanced diet (control) or the same balanced diet supplemented with 3 g fructose · kg(-1) · d(-1) and 6.77 g of a mixture of 5 essential amino acids 3 times/d (leucine, isoleucine, valine, lysine, and threonine) (HFrAA) or with 3 g fructose · kg(-1) · d(-1) and a maltodextrin placebo 3 times/d (HFr); there was a washout period of 4 to 10 wk between each condition. For each condition, the intrahepatocellular lipid (IHCL) concentration, VLDL-triglyceride concentration, and VLDL-[(13)C]palmitate production were measured after oral loading with [(13)C]fructose. RESULTS: HFr increased the IHCL content (1.27 ± 0.31 compared with 2.74 ± 0.55 vol %; P < 0.05) and VLDL-triglyceride (0.55 ± 0.06 compared with 1.40 ± 0.15 mmol/L; P < 0.05). HFr also enhanced VLDL-[(13)C]palmitate production. HFrAA significantly decreased IHCL compared with HFr (to 2.30 ± 0.43 vol%; P < 0.05) but did not change VLDL-triglyceride concentrations or VLDL-[(13)C]palmitate production. CONCLUSIONS: Supplementation with essential amino acids blunts the fructose-induced increase in IHCL but not hypertriglyceridemia. This is not because of inhibition of VLDL-[(13)C]palmitate production. This trial was registered at www.clinicaltrials.gov as NCT01119989.

Role of dietary carbohydrates and macronutrients in the pathogenesis of nonalcoholic fatty liver disease.

PURPOSE OF REVIEW: The prevalence of nonalcoholic fatty liver disease is increasing worldwide and there is strong evidence that dietary factors play a role in its pathogenesis. The present review aims to provide a better understanding of how carbohydrates and other macronutrients may affect the disease. RECENT FINDINGS: The effects of carbohydrates on the development of nonalcoholic fatty liver disease differ depending upon the carbohydrate type; high-glycemic index foods are related to increased hepatic fat in both rodents and humans. Similarly, simple carbohydrates, such as fructose, stimulate hepatic de-novo lipogenesis and decrease lipid oxidation, thus leading to increased fat deposition. The underlying mechanisms may involve the activation of transcription factors. Fat intake broadly leads to hepatic fat deposition in rodents but few data are available on humans. Both carbohydrates and fat trigger inflammatory factors, which are closely related to metabolic disorders and nonalcoholic fatty liver disease. Lifestyle interventions appear to be the most appropriate first-line treatment for nonalcoholic fatty liver disease. SUMMARY: There is strong evidence that the diet may affect the development of nonalcoholic fatty liver disease. Although simple carbohydrates are clearly shown to have deleterious effects in humans, the role of fat remains controversial. Further studies will be required to evaluate the effects of macronutrient composition on the development of nonalcoholic fatty liver disease.

Handling macromolecule signals in the quantification of the neurochemical profile.

In vivo localized proton magnetic resonance spectroscopy (1H MRS) became a powerful and unique technique to non-invasively investigate brain metabolism of rodents and humans. The main goal of 1H MRS is the reliable quantification of concentrations of metabolites (neurochemical profile) in a well-defined region of the brain. The availability of very high magnetic field strengths combined with the possibility of acquiring spectra at very short echo time have dramatically increased the number of constituents of the neurochemical profile. The quantification of spectra measured at short echo times is complicated by the presence of macromolecule signals of particular importance at high magnetic fields. An error in the macromolecule estimation can lead to substantial errors in the obtained neurochemical profile. The purpose of the present review is to overview methods of high field 1H MRS with a focus on the metabolite quantification, in particular in handling signals of macromolecules. Three main approaches of handling signals of macromolecules are described, namely mathematical estimation of macromolecules, measurement of macromolecules in vivo, and direct acquisition of the in vivo spectrum without the contribution of macromolecules.

Morphological and physiological species-dependent characteristics of the rodent Grueneberg ganglion.

In the mouse, the Grueneberg ganglion (GG) is an olfactory subsystem implicated both in chemo- and thermo-sensing. It is specifically involved in the recognition of volatile danger cues such as alarm pheromones and structurally-related predator scents. No evidence for these GG sensory functions has been reported yet in other rodent species. In this study, we used a combination of histological and physiological techniques to verify the presence of a GG and investigate its function in the rat, hamster, and gerbil comparing with the mouse. By scanning electron microscopy (SEM) and transmitted electron microscopy (TEM), we found isolated or groups of large GG cells of different shapes that in spite of their gross anatomical similarities, display important structural differences between species. We performed a comparative and morphological study focusing on the conserved olfactory features of these cells. We found fine ciliary processes, mostly wrapped in ensheating glial cells, in variable number of clusters deeply invaginated in the neuronal soma. Interestingly, the glial wrapping, the amount of microtubules and their distribution in the ciliary processes were different between rodents. Using immunohistochemistry, we were able to detect the expression of known GG proteins, such as the membrane guanylyl cyclase G and the cyclic nucleotide-gated channel A3. Both the expression and the subcellular localization of these signaling proteins were found to be species-dependent. Calcium imaging experiments on acute tissue slice preparations from rodent GG demonstrated that the chemo- and thermo-evoked neuronal responses were different between species. Thus, GG neurons from mice and rats displayed both chemo- and thermo-sensing, while hamsters and gerbils showed profound differences in their sensitivities. We suggest that the integrative comparison between the structural morphologies, the sensory properties, and the ethological contexts supports species-dependent GG features prompted by the environmental pressure.

Image-Derived Input Function from the Vena Cava for 18F-FDG PET Studies in Rats and Mice.

Measurement of arterial input function is a restrictive aspect for quantitative (18)F-FDG PET studies in rodents because of their small total blood volume and the related difficulties in withdrawing blood.

Assessment of terrestrial small mammals and a record of the critically endangered shrew Crocidura wimmeri in Banco National Park (Cote d'Ivoire)

This study investigated the small mammal community of the periurban Banco National Park (34 km(2)), Abidjan, Cote d'Ivoire, using identical numbers of Sherman and Longworth traps. We aimed to determine the diversity and distribution of rodents and shrews in three different habitats: primary forest, secondary forest and swamp. Using 5014 trap-nights, 91 individuals were captured that comprised seven rodent and four shrew species. The trapping success was significantly different for each species, i.e., the Longworth traps captured more soricids (31/36 shrews), whereas the Sherman traps captured more murids (37/55 mice). The most frequent species was Praomys cf. rostratus, followed by Crocidura buettikoferi, Hybomys trivirgatus and Crocidura jouvenetae. Indices of species richness (S) and diversity (H') were greatest in primary forest, followed by secondary forest and swamp. - Several expected species, such as Crocidura obscurior, were not found, whereas we captured four specimens of the critically endangered (IUCN 2012) Wimmer's shrew Crocidura wimmeri, a species that has vanished from its type locality, Adiopodoume. Therefore, Banco National Park represents an important sanctuary, not only for plants, birds and primates, but also for other small forest vertebrates.

PPAR expression and function during vertebrate development.

The peroxisome proliferator activated receptors (PPARs) are ligand activated receptors which belong to the nuclear hormone receptor family. As with other members of this superfamily, it is thought that the ability of PPAR to bind to a ligand was acquired during metazoan evolution. Three different PPAR isotypes (PPARalpha, PPARbeta, also called 6, and PPARgamma) have been identified in various species. Upon binding to an activator, these receptors stimulate the expression of target genes implicated in important metabolic pathways. The present article is a review of PPAR expression and involvement in some aspects of Xenopus laevis and rodent embryonic development. PPARalpha and beta are ubiquitously expressed in Xenopus early embryos but become more tissue restricted later in development. In rodents, PPARalpha, PPARbeta and PPARgamma show specific time- and tissue-dependent patterns of expression during fetal development and in the adult animals. PPARs are implicated in several aspects of tissue differentiation and rodent development, such as differentiation of the adipose tissue, brain, placenta and skin. Particular attention is given to studies undertaken by us and others on the implication of PPARalpha and beta in rodent epidermal differentiation.

Anti-Nogo-A Antibody Treatment Promotes Recovery of Manual Dexterity after Unilateral Cervical Lesion in Adult Primates--re-examination and Extension of Behavioral Data.

In rodents and nonhuman primates subjected to spinal cord lesion, neutralizing the neurite growth inhibitor Nogo-A has been shown to promote regenerative axonal sprouting and functional recovery. The goal of the present report was to re-examine the data on the recovery of the primate manual dexterity using refined behavioral analyses and further statistical assessments, representing secondary outcome measures from the same manual dexterity test. Thirteen adult monkeys were studied; seven received an anti-Nogo-A antibody whereas a control antibody was infused into the other monkeys. Monkeys were trained to perform the modified Brinkman board task requiring opposition of index finger and thumb to grasp food pellets placed in vertically and horizontally oriented slots. Two parameters were quantified before and following spinal cord injury: (i) the standard 'score' as defined by the number of pellets retrieved within 30 s from the two types of slots; (ii) the newly introduced 'contact time' as defined by the duration of digit contact with the food pellet before successful retrieval. After lesion the hand was severely impaired in all monkeys; this was followed by progressive functional recovery. Remarkably, anti-Nogo-A antibody-treated monkeys recovered faster and significantly better than control antibody-treated monkeys, considering both the score for vertical and horizontal slots (Mann-Whitney test: P = 0.05 and 0.035, respectively) and the contact time (P = 0.008 and 0.005, respectively). Detailed analysis of the lesions excluded the possibility that this conclusion may have been caused by differences in lesion properties between the two groups of monkeys.

Sex and clock to discover new PPARα functions

Summary : PPARα is a ligand-activated transcription factor that is a member of the nuclear receptor superfamily. In rodents, PPARα is highly expressed in liver, especially in parenchymal cells, where it has an impact on several hepatic functions such as nutrient metabolism, inflammation and metabolic stress. Ligands for PPARα comprise long chain unsaturated fatty acids, eicosanoids and lipid lowering fibrate drugs. In liver, many metabolic processes are orchestrated by the hepatic circadian clock. The aim of the hepatic clock is to synchronize cellular pathways allowing animals to adapt their metabolism to predictable daily changes in the environment. Indeed, similar to PPARα, the hepatic clock influences nutrient metabolism and detoxification through circadian output regulators :the PAR-domain basic leucine zipper proteins called PAR blip proteins. In this report, we showed that through a positive feedback loop mechanism, PAR. blip, proteins participate to the availability of PPARα endogenous ligands that contribute to the circadian expression and functions of PPARα. Interestingly, we also discovered some unexpected hepatic sexual dimorphic functions of PPARα. These functions are determined b PPARα sumoylation, interaction with DNA methylation mechanism and with unexpected proteins with gender specificity. The connection between circadian clock and hepatic sexual dimorphism opens new perspectives regarding the chronobiology of PPARα activity and the beneficial effects of PPARα agonist in the treatment of diseases related to steroid hormones metabolism characterized by inflammation and hepatotoxicity. Résumé : PPARα est un facteur de transcription activé par un ligand, membre de la superfamille des récepteurs nucléaires. Chez les rongeurs, PPARα est fortement exprimé dans le foie, spécialement dans les cellules du parenchyme dans lesquelles il joue un role important dans les fonctions hépatiques tels que le métabolisme des nutriments, l'inflammation et les stress métaboliques. Les ligands pour PPARα comprennent les acides gras à longues chaînes, les eicosanoides et les médicaments hypolipidémiques (fibrates). Dans le foie, beaucoup de processus métaboliques sont orchestrés par l'horloge circadienne hépatique. Le but de cette horloge est de synchroniser les voies métaboliqués permettant aux animaux d'adapter leurs métabolismes aux changements journaliers. Ainsi, l'horloge hépatique influence le métabolisme des nutriments tels que l'utilisation des lipides à travers certains régulateurs circadians appelés facteurs de transcription PAR bZips. Dans ce mémoire, nous avons montré qu'à travers une boucle de régulation, les protéines PAR bZip contrôlent la production des ligands endogènes à PPARα, jouant un rôle dans l'expression circadienne et les fonctions de PPARα. Nous avons également découvert des aspects méconnus des fonctions liées au dimorphisme sexuel de PPARα. Nous avons montré que PPARα est différemment sumoylisé entre les sexes et interagit avec la méthylation de l'ADN ainsi qu'avec des protéines insoupçonnées comme partenaires de PPARα. De part leur lien avec l'horloge circadienne et le dimorphisme sexuel, nos découvertes ouvrent de nouvelles perspectives concernant la chronobiologie de l'activité de PPARα et les effets bénéfiques des ses activateurs dans le traitement des maladies liées au métabolisme des hormones stéroides.

Chlamydiaceae and Chlamydia-like organisms in free-living small mammals in Europe and Afghanistan.

Few data are available on the occurrence of chlamydial infections in wild small mammals. We investigated the significance of free-living small mammals as reservoirs or transmission hosts for microorganisms of the phylum/class Chlamydiae. We obtained 3,664 tissue samples from 911 animals in Switzerland, Germany, Austria, the Czech Republic, and Afghanistan. Samples included internal organs (n = 3,652) and feces (n = 12) from 679 rodents (order Rodentia) and 232 insectivores (order Soricomorpha) and were tested by three TaqMan® real-time PCRs specific for members of the family Chlamydiaceae and selected Chlamydia-like organisms such as Parachlamydia spp. and Waddlia spp. Only one of 911 (0.11%) animals exhibited a questionable positive result by Chlamydiaceae-specific real-time PCR. Five of 911 animals were positive by specific real-time PCR for Parachlamydia spp. but could not be confirmed by quantitative PCR targeting the Parachlamydia acanthamoebae secY gene (secY qPCR). One of 746 animals (0.13%) was positive by real-time PCR for Waddlia chondrophila. This result was confirmed by Waddlia secY qPCR. This is the first detection of Chlamydia-like organisms in small wildlife in Switzerland. Considering previous negative results for Chlamydiaceae in wild ruminant species from Switzerland, these data suggest that wild small mammals are unlikely to be important carriers or transport hosts for Chamydiaceae and Chlamydia-like organisms.

Covariation between plumage colour polymorphism and diet in the Barn Owl Tyto alba

Several hypotheses might explain the evolution and maintenance of colour morphs within animal populations. The 'alternative foraging strategy' hypothesis states that alternative colour morphs exploit different ecological niches. This hypothesis predicts that morphs differ in diet, either because foraging success on alternative prey species is morph-dependent or because differently coloured individuals exploit alternative habitats. I examined this prediction in the Barn Owl Tyto alba, a bird that varies in plumage coloration continuously from dark reddish-brown to white. On the European continent, Owls are light-coloured (subspecies T. a. alba) in the south and reddish-brown (T. a. guttata) in the north; in central Europe the two subspecies interbreed, generating many colour variants. If plumage coloration indicates alternative foraging strategies, in sympatry dark- and light-coloured owls should consume prey species that are typical of the diets of T. a. guttata and T. a. alba in allopatry, respectively. In line with this prediction, both in allopatry and in sympatry in Switzerland T. a. guttata fed primarily upon Common Voles Microtus arvalis and T. a. alba upon Wood Mice Apodemus spp. Statistical analyses suggest that morph-dependent diet did not arise from a non-random habitat distribution of owls with respect to plumage coloration. This suggests that foraging success upon alternative prey is morph-dependent.

Asymmetric and differential gene introgression at a contact zone between two highly divergent lineages of field voles (Microtus agrestis).

Secondary contact zones have the potential to shed light on the mode and rate at which reproductive isolation accumulates during allopatric speciation. We investigated the population genetics of a contact zone between two highly divergent lineages of field voles (Microtus agrestis) in the Swiss Jura mountains. To shed light on the processes underlying introgression, we used maternally, paternally, and bi-parentally inherited markers. Though the two lineages maintained a strong genetic structure, we found some hybrids and evidence of gene flow. The extent of introgression varied with the mode of inheritance, being highest for mtDNA and absent for the Y chromosome. In addition, introgression was asymmetric, occurring only from the Northern to the Southern lineage. Both patterns seem parsimoniously explained by neutral processes linked to differences in effective sizes and sex-biased dispersal rates. The lineage with lower effective population size was also the more introgressed, and the mode-of-inheritance effect correlated with the male-biased dispersal rate of microtine rodents. We cannot exclude, however, that Haldane's effect contributed to the latter, as we found a marginally significant deficit in males (the heterogametic sex) among hybrids. We propose a possible demographic scenario to account for the patterns documented, and empirical extensions to further investigate this contact zone.

Hypoxie-ischémie cérébrale néonatale : rôle de la voie de JNK et implication de l'autophagie dans la mort neuronale

Résumé : Malgré les immenses progrès réalisés depuis plusieurs années en médecine obstétricale ainsi qu'en réanimation néonatale et en recherche expérimentale, l'asphyxie périnatale, une situation de manque d'oxygène autour du moment de la naissance, reste une cause majeure de mortalité et de morbidité neurologique à long terme chez l'enfant (retard mental, paralysie cérébrale, épilepsie, problèmes d'apprentissages) sans toutefois de traitement pharmacologique réel. La nécessité de développer de nouvelles stratégies thérapeutiques pour les complications de l'asphyxie périnatale est donc aujourd'hui encore essentielle. Le but général de ce travail est l'identification de nouvelles cibles thérapeutiques impliquées dans des mécanismes moléculaires pathologiques induits par l'hypoxie-ischémie (HI) dans le cerveau immature. Pour cela, le modèle d'asphyxie périnatale (proche du terme) le plus reconnu chez le rongeur a été développé (modèle de Rice et Vannucci). Il consiste en la ligature permanente d'une artère carotide commune (ischémie) chez le raton de 7 jours combinée à une période d'hypoxie à 8% d'oxygène. Il permet ainsi d'étudier les lésions de type hypoxique-ischémique dans différentes régions cérébrales dont le cortex, l'hippocampe, le striatum et le thalamus. La première partie de ce travail a abordé le rôle de deux voies de MAPK, JNK et p38, après HI néonatale chez le raton à l'aide de peptides inhibiteurs. Tout d'abord, nous avons démontré que D-JNKI1, un peptide inhibiteur de la voie de JNK présentant de fortes propriétés neuroprotectrices dans des modèles d'ischémie cérébrale adulte ainsi que chez le jeune raton, peut intervenir sur différentes voies de mort dont l'activation des calpaïnes (marqueur de la nécrose précoce), l'activation de la caspase-3 (marqueur de l'apoptose) et l'expression de LC3-II (marqueur de macroautophagie). Malgré ces effets positifs le traitement au D-JNKI1 ne modifie pas l'étendue de la lésion cérébrale. L'action limitée de D-JNKI1 peut s'expliquer par une implication modérée des JNKs (faiblement activées et principalement l'isotype JNK3) après HI néonatale sévère. Au contraire, l'inhibition de la voie de nNOS/p38 par le peptide DTAT-GESV permet une augmentation de 20% du volume du tissu sain à court et long terme. Le second projet a étudié les effets de l'HI néonatale sur l'autophagie neuronale. En effet, l'autophagie est un processus catabolique essentiel au bien-être de la cellule. Le type principal d'autophagie (« macroautophagie » , que nous appellerons par la suite « autophagie ») consiste en la séquestration d'éléments à dégrader (protéines ou organelles déficients) dans un compartiment spécialisé, l'autophagosome, qui fusionne avec un lysosome pour former un autolysosome où le contenu est dégradé par les hydrolases lysosomales. Depuis peu, l'excès ou la dérégulation de l'autoptiagie a pu être impliqué dans la mort cellulaire en certaines conditions de stress. Ce travail démontre que l'HI néonatale chez le raton active fortement le flux autophagique, c'est-à-dire augmente la formation des autophagosomes et des autolysosomes, dans les neurones en souffrance. De plus, la relation entre l'autophagie et l'apoptose varie selon la région cérébrale. En effet, alors que dans le cortex les neurones en voie de mort présentent des caractéristiques mixtes apoptotiques et autophagiques, ceux du CA3 sont essentiellement autophagiques et ceux du CA1 sont principalement apoptotiques. L'induction de l'autophagie après HI néonatale semble donc participer à la mort neuronale soit par l'enclenchement de l'apoptose soit comme mécanisme de mort en soi. Afin de comprendre la relation pouvant exister entre autophagie et apoptase un troisième projet a été réalisé sur des cultures primaires de neurones corticaux exposés à un stimulus apoptotique classique, la staurosporine (STS). Nous avons démontré que l'apoptose induite par la STS était précédée et accompagnée par une forte activation du flux autophagique neuronal. L'inhibition de l'autophagie de manière pharmacologique (3-MA) ou plus spécifiquement par ARNs d'interférence dirigés contre deux protéines autophagiques importantes (Atg7 et Atg5) a permis de mettre en évidence des rôles multiples de l'autophagie dans la mort neuronale. En effet, l'autophagie prend non seulement part à une voie de mort parallèle à l'apoptose pouvant être impliquée dans l'activation des calpaïnes, mais est également partiellement responsable de l'induction des voies apoptotiques (activation de la caspase-3 et translocation nucléaire d'AIF). En conclusion, ce travail a montré que l'inhibition de JNK par D-JNKI1 n'est pas un outil neuroprotecteur efficace pour diminuer la mort neuronale provoquée par l'asphyxie périnatalé sévère, et met en lumière deux autres voies thérapeutiques beaucoup plus prometteuses, l'inhibition de nNOS/p38 ou de l'autophagie. ABSTRACT : Despite enormous progress over the last«decades in obstetrical and neonatal medicine and experimental research, perinatal asphyxia, a situation of lack of oxygen around the time of the birth, remains a major cause of mortality and long term neurological morbidity in children (mental retardation, cerebral palsy, epilepsy, learning difficulties) without any effective treatment. It is therefore essential to develop new therapeutic strategies for the complications of perinatal asphyxia. The overall aim of this work was to identify new therapeutic targets involved in pathological molecular mechanisms induced by hypoxia-ischemia (HI) in the immature brain. For this purpose, the most relevant model of perinatal asphyxia (near term) in rodents has been developed (model of Rice and Vannucci). It consists in the permanent ligation of one common carotid artery (ischemia) in the 7-day-old rat combined with a period of hypoxia at 8% oxygen. This model allows the study of the hypoxic-ischemic lesion in different brain regions including the cortex, hippocampus, striatum and thalamus. The first part of this work addressed the role of two MAPK pathways (JNK and p38) after rat neonatal HI using inhibitory peptides. First, we demonstrated that D-JNKI1, a JNK peptide inhibitor presenting strong neuroprotective properties in models of cerebral ischemia in adult and young rats, could affect different cell death mechanisms including the activation of calpain (a marker of necrosis) and caspase-3 (a marker of apoptosis), and the expression of LC3-II (a marker of macroautophagy). Despite these positive effects, D-JNKI1 did not modify the extent of brain damage. The limited action of D-JNKI1 can be explained by the fact that JNKs were only moderately involved (weakly activated and principally the JNK3 isotype) after severe neonatal HI. In contrast, inhibition of nNOS/p38 by the peptide D-TAT-GESV increased the surviving tissue volume by around 20% at short and long term. The second project investigated the effects of neonatal HI on neuronal autophagy. Indeed, autophagy is a catabolic process essential to the well-being of the cell. The principal type of autophagy ("macroautophagy", that we shall henceforth call "autophagy") involves the sequestration of elements to be degraded (deficient proteins or organelles) in a specialized compartment, the autophagosome, which fuses with a lysosome to form an autolysosome where the content is degraded by lysosomal hydrolases. Recently, an excess or deregulation of autophagy has been implicated in cell death in some stress conditions. The present study demonstrated that rat neonatal HI highly enhanced autophagic flux, i.e. increased autophagosome and autolysosome formation, in stressed neurons. Moreover, the relationship between autophagy and apoptosis varies according to the brain region. Indeed, whereas dying neurons in the cortex exhibited mixed features of apoptosis and autophagy, those in CA3 were primarily autophagíc and those in CA1 were mainly apoptotic. The induction of autophagy after neonatal HI seems to participate in neuronal death either by triggering apoptosis or as a death mechanism per se. To understand the relationships that may exist between autophagy and apoptosis, a third project has been conducted using primary cortical neuronal cultures exposed to a classical apoptotic stimulus, staurosporine (STS). We demonstrated that STS-induced apoptosis was preceded and accompanied by a strong activation of neuronal autophagic flux. Inhibition of autophagy pharmacologically (3-MA) or more specifically by RNA interference directed against two important autophagic proteins (Atg7 and AtgS) showed multiple roles of autophagy in neuronal death. Indeed, autophagy was not only involved in a death pathway parallel to apoptosis possibly involved in the activation of calpains, but was also partially responsible for the induction of apoptotic pathways (caspase-3 activation and AIF nuclear translocation). In conclusion, this study showed that JNK inhibition by D-JNKI1 is not an effective neuroprotective tool for decreasing neuronal death following severe perinatal asphyxia, but highlighted two more promising therapeutic approaches, inhibition of the nNOSlp38 pathway or of autophagy.

Dexamethasone induces posttranslational degradation of GLUT2 and inhibition of insulin secretion in isolated pancreatic beta cells. Comparison with the effects of fatty acids.

GLUT2 expression is strongly decreased in glucose-unresponsive pancreatic beta cells of diabetic rodents. This decreased expression is due to circulating factors distinct from insulin or glucose. Here we evaluated the effect of palmitic acid and the synthetic glucocorticoid dexamethasone on GLUT2 expression by in vitro cultured rat pancreatic islets. Palmitic acid induced a 40% decrease in GLUT2 mRNA levels with, however, no consistent effect on protein expression. Dexamethasone, in contrast, had no effect on GLUT2 mRNA, but decreased GLUT2 protein by about 65%. The effect of dexamethasone was more pronounced at high glucose concentrations and was inhibited by the glucocorticoid antagonist RU-486. Biosynthetic labeling experiments revealed that GLUT2 translation rate was only minimally affected by dexamethasone, but that its half-life was decreased by 50%, indicating that glucocorticoids activated a posttranslational degradation mechanism. This degradation mechanism was not affecting all membrane proteins, since the alpha subunit of the Na+/K+-ATPase was unaffected. Glucose-induced insulin secretion was strongly decreased by treatment with palmitic acid and/or dexamethasone. The insulin content was decreased ( approximately 55 percent) in the presence of palmitic acid, but increased ( approximately 180%) in the presence of dexamethasone. We conclude that a combination of elevated fatty acids and glucocorticoids can induce two common features observed in diabetic beta cells, decreased GLUT2 expression, and loss of glucose-induced insulin secretion.