The aquatic environment is severely impacted by xenobiotics that are released due to anthropogenic activities, threatening ecosystem health. Some of these contaminants accumulate in lipophilic fish tissues and are maternally transferred to developing offspring, affecting their growth and performance. However, knowledge about the long-term and generational impacts associated with maternal transfer of contaminants is limited in fish. In this thesis, the hypothesis tested was that maternal transfer of bisphenol A (BPA) leads to disruption in the developmental programing of growth and stress axes functioning in rainbow trout (Oncorhynchus mykiss), and that these changes are passed on to the next generation. This was tested by exposing oocytes to either control (vehicle; <0.01% ethanol) 0.3, 3.0, and 30.0 mg l-1 BPA in ovarian fluid for 3 h, prior to fertilization, to mimic maternal transfer. This led to the accumulation of 0, 0.8, 4.4 and 41.3 ng BPA embryo-1. Oocytes were fertilized with milt from clean males, and offspring growth, development and stress performances were assessed in a clean environment for a year (F1 generation). For F2 generation, oocytes collected from F1 females, raised from the different BPA accumulated eggs, were fertilized with milt from clean males and raised in a clean environment for one year as described for F1 generation. The accumulated BPA in eggs was quickly cleared and it was no longer detected in the F1 embryos at hatch. BPA exposure reduced specific growth rate and increased food conversion ratio in larvae reared from BPA-laden oocytes. Moreover, BPA-exposed fish had an altered cortisol developmental profile and a delay in stress axis maturation. In addition, the mRNA abundance of genes involved in somatotropic [insulin-like growth factor (IGF) -1; IGF-2; IGF receptor b (IGF-1rb)] and stress axes functioning [steroidogenic acute regulatory protein (StAR); cytochrome P450 side chain cleavage (P450scc)] were altered. Also, changes in thyroid signaling [thyroid receptor (TR) mRNA levels] and cortisol signaling [glucocorticoid receptor (GR) protein expression] were disrupted temporally during development. These results demonstrate that BPA accumulation in eggs, mimicking maternal transfer, impacts growth and development, and delays stress axis maturation via non-reproductive endocrine disrupting routes in trout. Some of the BPA changes seen in F1 generation also persisted in the F2 generation. For instance, ancestral exposure to BPA led to reduced growth and whole body glycogen content prior to feeding in the F2 fish. The developmental transcript profile of growth hormone-1and -2, IGF-1 and -2 and IGF-1rb, along with whole body cortisol levels were impacted by ancestral exposure to BPA. Moreover, a delay in cortisol dynamics post-stress was noted in the F2 fish of BPA exposure lineage. Our results show that ancestral exposure to BPA leads to effects on growth and stress performance in rainbow trout, but the mechanism is not known. To further investigate the long-term effect of BPA accumulation in eggs on stress performances, F1 and F2 juvenile fish were subjected to an acute stressor. Also, head kidney tissues from these juvenile fish were subjected to adrenocorticotrophic hormone (ACTH) stimulation in vitro to assess cortisol production capacity. BPA accumulation in eggs led to a reduced acute handling stressor-induced plasma cortisol response in trout from the F1 and F2 (only high BPA group) generations. Also, BPA exposure had a pronounced impact on acute handling stressor-mediated plasma glucose (only F2 generation) and lactate levels, indicative of a metabolic disturbance. BPA exposure (only the 4.4 ng group) did affect unstimulated but not stimulated [ACTH or 8-bromo-cyclic AMP (8-B-cAMP)] cortisol production from head kidney slices of juvenile fish from F1 generation. In the F2 generation, there was an increase in ACTH-stimulated cortisol production only from the high BPA-exposed group. Overall, BPA in eggs disrupts long-term cortisol and metabolic stress performances in rainbow trout. While the impaired plasma cortisol stress performance was dose-related in the F1, the effect was apparent only for high BPA group in the F2 generation, suggesting that the generational effects on cortisol stress axis functioning may be concentration-dependent. A metabolomics approach further confirmed multigenerational effects associated with BPA accumulation in eggs. Analysis of the metabolome profile at hatch and prior to first feed, using gas chromatography-time of flight-mass spectrometry (GC-TOF-MS), revealed a BPA-mediated metabolic disruption, including changes in pathways involved in carbohydrate, lipid and amino sugar metabolism, and amino acid metabolism and synthesis. Pathways involved in citric acid cycle and alanine, aspartate and glutamate metabolism were altered in both generations, suggesting that these pathways have the potential to be markers with predictive value for multigenerational effects of BPA in fish. Altogether, the study provides novel insights on the impact of BPA on rainbow trout metabolome at hatch and first feed. The results suggest that pathways involved in energy metabolism are targets for BPA impact and should be investigated as potential markers for BPA toxicity. Overall, BPA accumulation in oocytes induces long-term delays in growth and stress axis maturation in F1 generations fish, and these effects persist in the F2 generation. The developmental profiles of key genes of the somatotropic and HPI axes were altered by BPA, along with whole body composition, suggesting that BPA exposure leads to a metabolic disturbance in fish, resulting in reduced growth. Additionally, the altered plasma cortisol response to acute stress in F1 and F2 juveniles provides evidence for multigenerational effects of BPA on stress axis functioning. The current study proposes that BPA-induced epigenetic modifications during early development may be playing a key role in the generational effects on growth and stress axes disruption in trout. The finding that the growth and developmental changes to BPA exposure also corresponds with endocrine and metabolome changes in multiple generations in trout is novel, and underscores the necessity to develop new risk assessments tools for chemicals that are maternally transferred in fish.

JAK/STAT signaling pathway is one of the most studied intracellular cascades transmitting signals from the extracellular environment to the cell nucleus in order to affect expression of target genes. Circadian clocks localized in the suprachiasmatic nuclei (SCN) of the hypothalamus are sensitive especially to light but they can respond to non-photic stimuli such as growth factors, opioids, leptin and cytokines that have been demonstrated to perform its function via the JAK/STAT signaling pathway. The recent findings of our laboratory demonstrated that STAT3 protein is highly produced by SCN of rat. Primary aim of our experiments was to test the circadian regulation of STAT3 production in SCN and describe the effect of exogenously administered leptin on STAT3 phosphorylation in the SCN, pineal gland and hypothalamic structures responsible for regulated feeding behavior and energy metabolism. Because activation of leptin receptors may stimulate a number of other signaling cascades, we chose phosphorylated forms of kinase ERK1/2 and GSK-3β as other markers of intracellular changes after administration of leptin in the studied structures. Our results proved rhythmic production of STAT3 protein in SCN of rat and indicated circadian regulation of sensitivity to leptin in hypothalamic structures. The data...

L'hypothalamus latéral (LHA) est impliqué dans un grand nombre de fonctions dont les principales sont la régulation du métabolisme énergétique et des états de vigilance. L'objectif de ce travail est de caractériser les grandes divisions du LHA et d'analyser leur implication dans le cycle veille / sommeil et la prise alimentaire. Une comparaison de la distribution de différents marqueurs hypothalamiques révélés par immunohistochimie ou hybridation in situ a permis de mettre en évidence cinq domaines dans le LHA. L'implication de chacun des domaines dans des conditions expérimentales liées aux états d'éveil ou à la prise alimentaire a été définie à partir de l'expression d'un marqueur d'activation neuronale, la protéine c-Fos. LeLHA antérieur et le LHA tubéral, chacun divisé en deux domaines rostral et caudal, sont composés de neurones localisés de manière diffuse et sont impliqués dans les phénomènes d'activation générale du système nerveux central. Le LHA mamillaire est renommé LHA prémamillaire à partir de l'étude de la distribution des ARNm GAD et préprotachykinine, et est composé de différents noyaux caractérisés par l'expression de marqueurs spécifiques.Parmi eux, les noyaux parasousthalamique (PSTN) et calbindine (CbN) sont activés lors d'exposition à des goûts de valeur hédonique positive. Les analyses hodologiques ont montré que ce complexe nucléaire reçoit des projections CGRP du noyau parabrachial et projette dansle noyau central de l'amygdale. Il est également connecté avec le cortex insulaire. Ces différentes structures sont impliquées dans les circuits de la gustation et de la récompense. Le PSTN et le CbN sont donc intégrés dans différentes voies de régulation du comportement de prise alimentaire. Enfin, les données morpho fonctionnelles obtenues pour le LHA sont mises en relation avec des réseaux impliquant diverses structures cérébrales telles que le télencéphale basal, le thalamus ou le mésencéphale. / The lateral hypothalamic area (LHA) is involved in many functions, but mainly in food intakeor sleep / wake cycle regulation. The aim of this study is to characterize the main regions inthe LHA and to analyze their involvement in the sleep / wake cycle and food intake. Acomparison of the distribution of different hypothalamic markers labeled byimmunohistochemistry or in situ hybridization highlights five domains in the LHA. Theimplication of each domain in different experimental conditions linked to arousal or foodintake is analyzed by studying the expression of the c-Fos protein as a neuronal activationmarker. Neurons in the anterior and tuberal LHA, both divided into rostral and caudaldomains, are distributed in a diffuse way and are activated under arousal conditions. Themammillary LHA, renamed the premammillary LHA on the basis of GAD andpreprotachykinin mRNA expression, is composed of several nuclei characterized by specificmarker expression. Among them, the parasubthalamic nucleus (PSTN) and the calbindinnucleus (CbN) are activated by ingestion of compounds with a positive taste. Theconnectional analysis showed that these two structures receive CGRP projections from theparabrachial nucleus and projects into the central nucleus of the amygdala. The PSTN / CbNcomplex is also connected with the insular cortex. These different structures are known to beinvolved in gustatory and reward circuitries; the PSTN and the CbN are thus integrated innetworks controlling the food intake behavior. Finally, the morphofunctional data obtainedfor the LHA demonstrate that this region is connected to other networks involving variouscerebral structures such as the basal telencephalon, the thalamus or the mesencephalon.

Stress reaction is usually activated by the brain, when homeostasis is or perceived to be threatened. The stress signals are transmitted from the brain by two main branches; the sympathoadrenomedullary and the hypothalamo-pituitary-adrenal (HPA) axes and employ neural, humoral and immune pathways to cope with the stressor. Because of its potency, the stress reaction has to be precisely regulated. The HPA axis is regulated by feedback loops where its end product, corticosterone in laboratory rat and mouse, inhibits its activity. The effect of corticosterone does not depend only on the concentration of corticosterone but also on local metabolism of glucocorticoids via oxo-reduction catalyzed by the enzyme 11β -hydroxysteroid dehydrogenase 1 (encoded by the Hsd11b1 gene), which intracellularly regenerates active corticosterone from inactive 11-dehydrocorticosterone, or by extra-adrenal de novo steroidogenesis of glucocorticoids. We focused on analysis of stress response in experimental animals differing in HPA axis responsivity (Fischer 344 rats (F344) vs. Lewis rats (LEW) and germ-free (GF) vs. specific pathogen free mice (SPF)) with special emphasis on regulation of stress response, glucocorticoid regeneration and influence of gut microbiota. We found that stress modulated local regeneration of...

The magnocellular neurosecretory cells (MNCs) of the hypothalamus project axons from the supraoptic nucleus to the posterior pituitary gland, where they secrete either oxytocin or vasopressin into the circulation. Oxytocin is important for delivery at birth and is essential for milk ejection during suckling. Vasopressin primarily promotes water reabsorption in the kidney to maintain body fluid balance. The profile of oxytocin and vasopressin secretion is principally determined by the pattern of action potentials initiated at the cell bodies in the hypothalamus. MNCs principally secrete hormones from terminals in the pituitary, but the secretion also occurs from their dendrites in the supraoptic nucleus, where they diffuse and affect the neighbouring cells. Mechanisms controlling the oxytocin and vasopressin secretion from MNCs have been extensively studied over the last decades and it is assumed that the relationship between Ca2+ signalling, secretion from dendrites, and the firing patterns is essential in understanding the magnocellular neurosecretory system. In this project, we combine mathematical analysis and experimental measurements of Ca2+ activity of MNCs of transgenic rats expressing an arginine vasopressin-enhanced green fluorescent protein (AVP-eGFP) or oxytocin-monomeric red fluorescent...

TBI stands for Traumatic Brain Injury and refers to damage to the brain resulting from an external physical force, such as a blow, jolt, or penetrating injury to the head. Common causes of TBI include falls, motor vehicle accidents, sports injuries, and violence and has been linked to thousands of deaths and injuries in the US and the EU alike. This thesis was aimed to localise certain TBI to a specific part of the brain by exerting similar loading conditions on an Finite Element Method (FEM) of the rat brain as physical experiments conducted on living rats. By comparing the strain in 7 vital parts of the brain to injury diagnosis conducted in the physical experiments, an effort was made to link localised strain to injury diagnosis. The results indicate that strain in the thalamus and hypothalamus are linked with a loss of consciousness while strain in the hypothalamus coupled with the neocortex correlates greatly with activity-based behaviour changes. Lastly, injury associated with emotional changes are believed to stem from large strains in the neocortex. There is a theory suggesting that the structure of myeline, which provides support in motion and movement patterns of biological systems in humans and animals (known as biomechanical kinematics), could have an impact. However, more studies are needed to confirm and determine the exact cause. / TBI, från engelskans Traumatic Brain Injury, står för Traumatisk Hjärn Skada och syftar på en skada i hjärnan till följd av enyttre fysisk kraft, såsom ett slag, stöt eller genomträngande skada i huvudet. Vanliga orsaker till TBI inkluderar fall, motorfordonsolyckor, sportskador och våld och har kopplats till tusentals dödsfall och skadade i både USA och EU. Denna rapport syftar till att försöka lokalisera viss TBI till en specifik del av hjärnan genom att utöva liknandebelastningsförhållanden på en finit elementmetod (FEM) modell av råtthjärnan som fysiska experimentutförs på levande råttor. Genom att jämföra belastningen i 7 vitala delar av hjärnan med skadediagnos som utfördes i de fysiska experimenten gjordes en ansträngning för att koppla lokaliserad belastning till skadediagnos. Resultaten indikerar att skada i thalamus och hypotalamus är kopplade till en förlust av medvetande medan belastning i hypotalamus i kombination med neocortex korrelerar kraftigtmed aktivitetsbaserade beteendeförändringar. Slutligen är skador i samband med känslomässiga förändringartros härröra från skada i neocortex. Det finns teori som tyder på attstruktur av myelin, som ger stöd i rörelse och rörelsemönster av biologiskasystem hos människor och djur (känd som biomekanisk kinematik), kan ha en inverkan.Det behövs dock fler studier för att bekräfta och fastställa den exakta orsaken.