La réduction du temps de sommeil est un phénomène de plus en plus courant. Un faisceau de données expérimentales et épidémiologiques suggère qu'un manque de sommeil pourrait être un facteur de risque d'obésité et de diabète. Dans un premier temps, puisque des modifications de l'axe Hypothalamo-Hypophyso-Surrénalien (HHS) pourraient sous- tendre la relation entre le manque de sommeil et les maladies métaboliques, j'ai évalué les effets de 2 nuits courtes sur ce système chez des jeunes hommes en bonne santé. Nous avons montré que 2 nuits de 4h au lit altéraient l'activité spontanée et la réactivité de l'axe HHS et que l'ampleur des altérations était corrélée à l'importance de la privation de sommeil. Dans un deuxième temps, j'ai tenté de déterminer si une extension de sommeil pouvait avoir des effets bénéfiques chez des jeunes obèses dormant habituellement peu. Nos résultats préliminaires montrent que par simple extension du temps passé au lit, des obèses dormant habituellement 6h, étaient capables de dormir 8h, une durée associée au plus faible risque d'obésité dans les études épidémiologiques, que leur appétit pour les aliments gras et salés et le grignotage diminuaient, que leurs taux de polypeptide pancréatique, une hormone anorexigène, étaient augmentés, et que leur prise calorique lors d'un buffet à volonté était d'autant plus diminuée que leur temps de sommeil était augmenté. Ce travail souligne l'importance d'une durée de sommeil suffisante pour une bonne santé métabolique et suggère que l'optimisation du sommeil pourrait constituer une alternative peu coûteuse pour la prévention et la prise en charge des maladies métaboliques / Voluntary sleep restriction is increasingly common in modern socities. Evidence from epidemiological and experimental studies suggest that sleep loss may be a risk factor for obesity and type 2 diabetes. First, since the modifications in Hypothalamic-Pituitary-adrenal (HPA) axis activity may underlie the relationship between sleep loss and metabolic diseases, 1 evaluated the effect of 2 short nights on this system in healthy lean young men. We showed that 2 nights of 4h in bed impaired spontaneous activity and the reactivity of the HPA axis and that the magnitude of these alterations was related of the severity of sleep loss. 1n a second step, 1 sought to determine if sleep extension could have a beneficial effect in young obese short sleepers. Our preliminary results showed that, by a simple bedtime extension, obese subjects usually sleeping 6h were able to sleep 8h, a duration associated with the lowest risk obesity risk in epidemiological studies. Moreover, their appetite for sweets and fat food, and snacking were decreased, the levels of pancreatic polypeptide, an anorexigenic hormone, were increased and the more they slept, the less they consumed calories at an ad libitum buffet. This work highlights the importance of getting enough sleep to maintain a good metabolic health and suggest that sleep optimization may have implications for novel public health interventions

abstract: Sleep is essential for physical and psychological health. Sleep has also been linked to the daily patterns of key stress-responsive physiological systems, specifically the hypothalamic-pituitary-adrenal (HPA) axis and autonomic nervous system (ANS). Extant research examining sleep and diurnal patterns of cortisol, the primary end product of the HPA axis, is inconsistent. Moreover, it is not clear how specific aspects of sleep behavior (e.g., sleep duration, sleep quality, sleep variability) are related to specific components of diurnal cortisol rhythms. Salivary alpha-amylase (sAA) has been recognized as a surrogate marker of ANS activity, but limited research has explored relations between sleep and sAA diurnal rhythms. The current study utilized a modified ecological momentary assessment protocol to examine within- and between-person relations between multiple facets of sleep behavior using multiple methods (e.g., subjective report, actigraphy) and salivary cortisol and sAA. First year college students (N = 76) provided saliva samples and diary entries five times per day over the course of three days. Sleep was assessed via questionnaire, through daily diaries, and monitored objectively using actigraphy over a four day period. Between-person results revealed that shorter average sleep duration and greater sleep variability was related to lower levels of waking cortisol and flatter diurnal slopes across the day. Within-person results revealed that on nights when individuals slept for shorter durations than usual they also had lower levels of waking cortisol the next day. Sleep was not related to the cortisol awakening response (CAR) or diurnal patterns of sAA, in either between-person or within-person analyses. However, typical sleep behaviors measured via questionnaire were related to waking levels of sAA. Overall, this study provides a greater understanding of how multiple components of sleep, measured in naturalistic environments, is related to cortisol and sAA diurnal rhythms, and how day-to-day, within-person changes in sleep duration contribute to daily variations in cortisol. / Dissertation/Thesis / Masters Thesis Psychology 2015

Made available in DSpace on 2014-12-17T15:36:51Z (GMT). No. of bitstreams: 1 MarciaAFM.pdf: 1370878 bytes, checksum: 51aee5b7941400056788fa2e93d0df1a (MD5) Previous issue date: 2008-03-28 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The light, besides the vision stimuli, controls other process completely independent of image formation, such as the synchronization of the organismic circadian rhythms to the enviromental light/dark cycle. In mammals, this adjust occurs through the retinohypothalamic tract, a direct retinal projection to the suprachiasmatic nucleus, considered to be the major circadian pacemaker. Early studies have identified only the suprachiasmatic nucleus as a retinal target in the hypothalamus. However, using more sensitive neuroanatomic tracers, other retinorecipient hypothalamic regions outside to suprachiasmatic nucleus were pointed in a great number of mammalian species. In this study, the retinohypothalamic tract was shown in the rock cavy (Kerodon rupestris), an endemic rodent of the semiarid region of the Brazilian Northeast, using unilateral intravitreal injections of cholera toxin subunit b as a neuronal tracer. The results reveal that in the rock cavy, besides the suprachiasmatic nucleus, several hypothalamic regions receive direct retinal projection, such as the ventrolateral preoptic nucleus, medial and lateral preoptic areas, the supraoptic nucleus and bordering areas, anterior, lateral and rectrochiasmatic hypothalamic areas, and the subparaventricular zone. The results are discussed by comparing with those of the literature, into a functional context / A luz, al?m de ser o est?mulo para a vis?o, tamb?m controla processos que s?o completamente independentes da forma??o de imagem, como o ajuste dos ritmos circadianos do organismo ao ciclo claro/escuro ambiental. Em mam?feros, este ajuste ocorre atrav?s do trato retino-hipotal?mico, uma proje??o direta da retina para o n?cleo supraquiasm?tico, o principal marcapasso circadiano. Os trabalhos pioneiros mostraram a proje??o retino-hipotal?mica exclusivamente para o n?cleo supraquiasm?tico. Entretanto, atrav?s do uso de tra?adores neurais mais sens?veis, outras ?reas hipotal?micas retino-recipientes foram apontadas em um amplo n?mero de esp?cies de mam?feros. Neste trabalho o trato retino-hipotal?mico no moc? (Kerodon rupestris), um roedor end?mico da caatinga brasileira, foi demonstrado atrav?s de inje??o intra-ocular unilateral da subunidade b da toxina col?rica como tra?ador neuronal e revela??o imuno-histoqu?mica. Os nossos resultados mostram que, no moc?, al?m do n?cleo supraquiasm?tico, v?rias outras ?reas hipotal?micas recebem proje??o direta da retina, tais como o n?cleo pr?-?ptico ventrolateral, as ?reas pr?-?pticas medial e lateral, o n?cleo supra-?ptico e adjac?ncias, as ?reas hipotal?micas anterior, lateral e retroquiasm?tica e a zona subparaventricular. Os resultados s?o discutidos ? luz dos dados da literatura, dentro de um contexto funcional

Submitted by Rafael Santana (rafael.silvasantana@ufpe.br) on 2017-05-12T18:27:18Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação.pdf: 1932966 bytes, checksum: e5c87665e0d22baf18bc136471875d00 (MD5) / Made available in DSpace on 2017-05-12T18:27:18Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação.pdf: 1932966 bytes, checksum: e5c87665e0d22baf18bc136471875d00 (MD5) Previous issue date: 2016-03-04 / FACEPE / Diversos estudos evidenciaram os efeitos da desnutrição perinatal sobre alterações morfofisiológicas no organismo adulto. No presente estudo, foi verificado os efeitos da desnutrição proteica perinatal sobre a expressão de receptores 5-HT1B nos núcleos paraventricular (PVN) e arqueado do hipotálamo, e no Caudado-Putamem (CPu) e Amigdala Basolateral (ABL). Além disso, foi analisado ainda os efeitos que essa desnutrição promoveu sobre a ativação neural no CPu, ABL e na Amigdala Central (AC). Foram utilizados dois principais grupos de ratos da linhagem Wistar: um grupo submetido a desnutrição proteica (8% caseína) durante o período da gestação e lactação; e o outro com dieta padrão (17% caseína). No 35º dia de vida os animais foram subdivididos em quatro grupos: Desnutrido que recebeu solução salina (D), Desnutrido que recebeu d-fenfluramina (DF), Controle que recebeu solução salina (C) e Controle que recebeu d-fenfluramina (CF), nas proporções de 3mg/kg p.c para fenfluramina e 1ml/kg para solução salina. Observou-se maior número de células imunorreativas (p<0,01) ao 5-HT1B na ABL no grupo desnutrido (33,8 ± 10,05, n=4) comparado ao controle (5,3 ± 1,53, n=3), enquanto que no CPu não houve modificação. No hipotálamo, foi verificado aumento de células 5-HT1B no grupo desnutrido comparado ao controle nos núcleos ARC (C= 14,3 ± 6,66, n=3 versus D= 26,8± 4,19, n=4, p<0,01) e PVN (C= 30 ± 8,19, n=3 versus D= 58,3 ± 16,66, n=4, p<0,05). Após a administração de fenfluramina, houve aumento no número de neurônios imunorreativos a proteína FOS no grupo desnutrido fenfluramina comparado ao desnutrido salina no CPu (D= 26,5 ± 21,56, n=4 versus DF= 106,5 ± 40,45, n=4, p<0,05) e AC (D= 13,6 ± 8,17, n=5 versus DF= 64 ± 31,48, n=3, p<0,05). No grupo controle, a fenfluramina não alterou o número de células ativadas no CPu (C= 16,5 ± 7,78, n=2 versus CF= 63 ± 39,28, n=3, p>0,05) e AC (C= 6,3 ± 4,93, n=3 versus CF= 44 ± 26,33, n=5, p>0,05). Não houve alteração no número de células imunorreativas a proteína FOS na ABL no grupo controle (C= 5 ± 3,37, n=4 versus CF= 21,3 ± 11,35, n=4, p>0,05) ou desnutrido (D= 20 ± 13,64, n=4 versus DF= 14,4 ± 3,13, n=5, p>0,05) quando comparados aos seus respectivos grupos salina. Este estudo evidenciou que a desnutrição proteica no período perinatal acarreta aumento na expressão de células imunorreativas contra o receptores 5-HT1B nos núcleos ARC, PVN e ABL tal como aumenta a ativação de neurônios no CPu e na AC em resposta a fenfluramina. / Several studies have shown the effects of perinatal malnutrition on morphological and physiological changes in the adult organism. In the present study was investigated the effects of perinatal protein malnutrition on the expression of 5-HT1B receptors in the paraventricular nucleus (PVN) of the hypothalamus, arcuate, caudate-putamen (CPu) and basolateral amygdala (GLA). In addition, it was analyzed the effects that malnutrition promoted on the neural activation of the CPu, Central GLA and the amygdala (AC). Two main groups of rats Wistar were used: A group subjected to protein malnutrition (8% casein) during the period of pregnancy and lactation and the other with standard diet (17% casein). On the 35th day of life the animals were divided into four groups: Malnourished received saline (D) Malnourished that received dfenfluramine (DF), control which received saline (C) and control that received dfenfluramine (CF) in the proportions of 3 mg/kg of BW fenfluramine and 1 ml/kg for saline. There were a higher number of immunoreactive cells (p <0.01) to the 5-HT 1B in the ABL in the undernourished group (33.8 ± 10.05, n = 4) compared to the control (5.3 ± 1.53, n = 3), whereas in the CPu no change was observed. In the hypothalamus, it was observed increase of 5-HT 1B cells in the undernourished group compared to the control in ARC cores (C = 14.3 ± 6.66, n = 3 and D = 26.8 ± 4.19, n = 4, p <0.01) and the PVN (C = 30 ± 8.19, n = 3 and D = 58.3 ± 16.66 n = 4, p <0.05). After administration of fenfluramine, there was an increase in the number of immunoreactive neurons FOS protein in malnourished group fenfluramine compared to saline undernourished in CPu (D = 26.5 ± 21.56, n = 4 and DF = 106.5 ± 40.45 , n = 4, p <0.05) and AC (D = 13.6 ± 8.17, n = 5 and 31.48 ± DF = 64, n = 3, p <0.05). In the control group, fenfluramine did not change the number of activated cells in CPu (C = 16.5 ± 7.78, n = 2 and CF = 63 ± 39.28, n = 3, p> 0.05) and AC (C = 6.3 ± 4.93, n = 3 and CF = 44 ± 26.33, n = 5, p> 0.05). There was no change in the number of immunoreactive cells FOS protein in the ABL in the control group (C = 5 ± 3.37, n = 4 and CF = 21.3 ± 11.35, n = 4, p> 0.05) or malnourished (D = 20 ± 13.64, n = 4 versus DF = 14.4 ± 3.13, n = 5, p> 0.05) when compared to their saline groups. This study showed that the protein malnutrition in the perinatal period causes an increase in the expression of immunoreactive cells against the 5-HT 1B receptors in the ARC cores, PVN and ABL as well as increases the activation of neurons in the CPu and AC in response to fenfluramine.

A área incerto-hipotalâmica (IHy) é uma região diencefálica que atua em funções reprodutivas através da presença do hormônio concentrador de melanina (MCH). Nosso objetivo é estudar as conexões da IHy de fêmeas com áreas relacionadas ao controle neuroendócrino reprodutivo, usando traçador anterógrado dextrana amina biotinilada (BDA) e retrógrado Fluorogold (FG). Nesse sentido, destacamos que a IHy, em fêmeas, conecta-se reciprocamente com diversos núcleos hipotalâmicos da região periventricular e pré-óptica, além das áreas hipotalâmica anterior, posterior e lateral. Nosso resultados também sugerem um dimorfismo sexual das projeções da IHy, já que a área pré-óptica medial, o núcleo anteroventral periventrciular e o núcleo arqueado são mais densamente inervadas pelas IHy em fêmeas do que em machos. / The incerto-hypothalamic area (IHy) is a diencephalic region poorly studied which is characterized by two non-colocalized neurochemical groups composed by the A13 dopaminergic group intermingled with predominantly GABAergic cells co-expressing melanin-concentrating hormone (MCH) and/or cocaine and amphetamine regulated transcript (CART). Functional studies suggest that IHy is involved in the neuroendocrine control of female reproduction in specific metabolic states mediated by MCH. In this way, our aim is to study the IHy connections in female rats related to neuroendocrine control of female reproduction using neuronal anterograde Biotin Dextran Amine (BDA) and retrograde Flurogold (FG) tracers. As a result, we found that IHy projections are the lateral septal nucleus, bed nucleus of stria terminalis, paraventricular thalamic nucleus, reuniens thalamic nucleus, medial preoptic area, medial and lateral preoptic nucleus, median preoptic nucleus, periventricular hypothalamic nucleus, anteroventral periventricular nucleus, anterior, posterior and lateral hypothalamic area and precommissural nucleus. The main afferents to the IHy, in females, are the lateral septal nucleus, median preoptic nucleus and paraventricular thalamic nucleus. These results suggest sexually dimorphic projections from the IHy, since IHy more densely innervates neuroendocrine regions in female than in male rats.

A glândula pineal sintetiza o hormônio melatonina à noite e este regula diversos sistemas fisiológicos, adaptando-os às exigências de cada momento do dia. O objetivo deste trabalho foi o de analisar o perfil noturno da síntese de melatonina na glândula pineal de ratos, em diferentes idades, tratados com glutamato monossódico (MSG) no período neonatal. Ratos Wistar, machos e fêmeas, receberam injeções de MSG (4mg/g/dia) ou de solução salina (0,9%) do 2º ao 8º dia pós-natal. Foram avaliados o peso corporal e dos tecidos adiposos, o comprimento naso-anal,o perfil noturno da síntese de melatonina e da atividade da enzima AANAT, o GTT e o ITT. O MSG induziu um aumento no peso dos tecidos adiposos, sem aumento do peso corporal. Não foi observada hiperglicemia, nem intolerância à glicose, mas sim resistência insulínica. A ritmicidade circadiana da melatonina e da AANAT foi preservada, mas houve um aumento de ambos no ZT 15. As alterações na síntese de melatonina parecem decorrer das lesões hipotalâmicas provocadas pelo MSG, pela redução do NPY, aumento da insulina ou da noradrenalina. / The pineal gland synthesizes melatonin exclusively at night. Melatonin is a temporal synchronizer of several physiological functions, adapting the organism to the diurnal environmental changes. The purpose of this work was to analyze the effects of neonatal monosodium glutamate (MSG) administration on the nocturnal profile of melatonin synthesis in the pineal gland. Wistar rats, males and females, were injected neonatally with MSG (4mg/g/day) or saline solution (0.9%) from the 2nd to 8th post-natal day. Body weight and adipose tissue weight, naso-anal length, nocturnal melatonin and AANAT activity profiles, GTT and ITT were analyzed. MSG induced a great increase in adipose depots without increase in body weight. It did not induce hyperglycemia nor glucose intolerance, but induced insulin resistance. Circadian rhythmicity of melatonin synthesis and AANAT activity was not altered, but there was an increase at ZT 15 in both. It seems that melatonin synthesis changes could be related to hypothalamic lesions, through a reduction in NPY or insulin/norepinephrine elevation.

Glucocorticoid excess is associated with adverse effects on a number of physiological parameters, leading to obesity, dysfunction of the hypothalamic-pituitary- adrenal (HPA) axis and behavioural changes such as anxiety and impaired learning and memory. Circulating and local tissue glucocorticoid levels are tightly controlled by the HPA axis but an additional level of control exists in tissues such as brain, liver and adipose tissue. In these structures, enzymes including 5α-reductase 1 (5αR1), catalyse the conversion of corticosterone to A-ring reduced metabolites, which have a different spectrum of activities. This thesis investigates the role of 5αR1 in regulating central glucocorticoid actions which control HPA axis function and behaviour in a mouse model with genetic disruption of 5αR1 (5αR1-KO). Preliminary data showed 5αR1-KO mice were susceptible to developing insulin resistance and obesity and had reduced HPA axis responses to acute stress. Additionally, male 5αR1-KO mice were more prone to obesity than wild-type (WT) when fed a high-fat diet whilst female 5αR1-KO mice gained more weight than WT even on a normal chow diet. Intriguingly, female 5αR1-KO mice subjected to social isolation stress lost this extra weight and became comparable to WT controls. This study tested the hypothesis that 5αR1-KO mice are less able to inactivate glucocorticoids in the periphery and within tissues, resulting in a predisposition to metabolic disturbances and behavioural alterations. These were hypothesised to include hyperphagia, weight gain, impaired stress responses, anxiety (exacerbated by environmental stress) and cognitive deficits. It was also thought that many of these features would be more pronounced in female vs. male mice. The main aims of this study were to determine if 5αR1-KO induced weight gain and if this was correlated to altered gene expression of key hypothalamic neuropeptides which regulate appetite, to determine the central mechanisms which underpin attenuated HPA axis responses to acute stress and to determine whether behaviours such as anxiety and learning and memory ability are affected by global 5αR1 loss. It was hypothesised that female 5αR1-KO mice have increased appetite and reduced locomotor activity compared with WT and male 5αR1-KOs. However, male 5αR1- KO mice (on a mixed genetic background, C57Bl/6j/SvEv/129) were hyperphagic on a normal chow diet but did not gain extra weight, while female 5αR1-KO mice gained more weight vs. WT despite hypophagia. Free ambulatory activity was unaffected by genotype in either sex. Male 5αR1-KO mice appeared less anxious but responses of female 5αR1-KO mice in tests of anxiety did not differ from WT controls. Mice lacking 5αR1 generally had a poorer metabolic profile with impaired glucose tolerance and hyperinsulinaemia; with hepatic steatosis evident in female mice. There was evidence of compensatory changes in hypothalamic orexigenic and anorexigenic peptides. Phenotypes were sexually dimorphic such that male mice had a poorer metabolic profile vs. females, which was particularly marked in male 5αR1- KO animals. 5αR1-KO mice were previously shown to have attenuated HPA axis responses to acute stress and it was hypothesised that disruption of 5αR1 would result in altered expression of genes related HPA axis regulation with a view to increased negative feedback. Here, male and female 5αR1-KO mice demonstrated altered corticosteroid receptor expression within the hippocampus and the pituitary, two key structures in the HPA cascade. In situ hybridisation showed reduced mRNA for MR in the hippocampus and for Crh in the hypothalamus of 5αR1-KO mice. These modifications along with decreased Crhr-1 mRNA (CRH‘s main receptor) may be due to a lack of corticosterone metabolism within the brain resulting in enhanced negative feedback and reduced HPA axial drive. In order to study behaviour in detail and also to test whether potential central glucocorticoid excess may predispose to cognitive decline with ageing, a separate cohort of female 5αR1-KO backcrossed onto a uniform C57Bl/6j background was studied both when young (6 months) and when aged (14-15 months). Additionally, mice were housed in either groups or singly (social isolation) to investigate the potentially additive effects of environmental stress. It was hypothesised that local glucocorticoid increases in the brains of 5αR1-KO mice would be associated with anxiety and cognitive deficiencies and that these phenotypes would be exaggerated by the stress of social isolation as well as ageing. Behavioural differences were not observed at 6 months of age. However aged, 5αR1-KO mice housed singly showed increased anxiety and had higher plasma corticosterone levels than group-housed mice. Moreover, aged mice lacking 5αR1 performed less well than WT in tests of memory and had a marginally greater cognitive decline when learning ability at 14- 15 months old was compared to that of the same animals tested at 6 months old. Overall, mice with global 5αR1 loss appeared susceptible to anxiety as well as some degree of age-associated cognitive impairment, but only when subjected to social isolation stress which is a known chronic stressor. The final set of experiments aimed to determine the effect of mouse strain on 5αR1- KO phenotypes. It was hypothesised that glucocorticoid clearance would be attenuated to a lesser degree in 5αR1-KO mice bred onto a congenic C57Bl/6j strain compared to those of the mixed strain and that this would manifest as less disruption of metabolism and less suppression of HPA axis stress responses. Although social isolation again induced weight-loss in female mice and more so in 5αR1-KO animals, mice on the C57Bl/6j background strain did not show dampened HPA axis responses to acute stress as seen previously. It was subsequently shown in adrenalectomised mice that animals bred on the C57Bl/6j strain cleared active corticosterone from plasma and liver faster than mixed strain mice. This may have rendered mixed strain 5αR1-KO mice more susceptible to excessive corticosterone levels producing a more exaggerated phenotype in this group. In conclusion, these data suggest a role for the enzyme 5αR1 in modifying glucocorticoid concentrations in the brain and liver, influencing not only metabolic and peripheral effects such as weight gain and insulin resistance, but also in modifying cognition, appetite stimulation and affective behaviours. It has been highlighted that outside factors such as housing and age can modify these phenotypes and are important considerations for future studies. This study has also highlighted the importance of choosing an appropriate genetic background for genetically modified animals since phenotypes can be enhanced or attenuated depending on strain. Finally, 5αR inhibitors are used to treat disorders such as benign prostatic hyperplasia in men, and it is important to consider that these drugs may have a wide array of associated side effects both systemically and in the central nervous system.

The circadian timing system establishes daily rhythms in behavior and physiology throughout the body, ensuring that functions like activity, sleep and hormone release are appropriately timed. Research suggests that his temporal synchrony within the body is quite important for health and survival. In mammals, the central circadian pacemaker in the suprachiasmatic nucleus (SCN) drives rhythms in behavior and physiology in large part by stimulating or inhibiting other brain regions responsible for these functions at the appropriate times of day. This timed signal is often indirect, i.e. relayed or possibly processed through a series of neurons in different brain regions before reaching the effector site. The subparaventricular zone (SPZ), a region adjacent to the SCN which is the main recipient of direct neuronal inputs from the SCN, is thought to be a critical relay for SCN signals, since loss of the SPZ results in loss of circadian rhythms in body temperature, activity and sleep/wakefulness. Another important relay site, the dorsomedial hypothalamic nucleus (DMH) gets direct input from both the SCN and SPZ and is critical for normal expression of various circadian rhythms.

Epidemiological studies show that offspring subjected to maternal undernutrition during early pregnancy are prone to developing obesity and other diseases in adulthood. The hypothalamic energy regulating pathway may be altered in these offspring, with epigenetic changes as a core mechanism. Therefore, this thesis aimed to determine if epigenetic changes are present in this pathway in the hypothalami from offspring subjected to maternal undernutrition. The investigations are focused on the glucocorticoid receptor (GR) as an inhibitor of the anorexigenic neuropeptide pro-opiomelanocortin (POMC), with potential modifications leading to increased food intake and the development of obesity. To achieve this, an established sheep model developed by our collaborators was used, during which maternal ewes were undernourished periconceptionally to produce a 10-15% decrease in body weight. We found that hypothalami from fetal offspring had greater epigenetic modifications when this reduction in maternal body weight was maintained from 60 days before conception until 30 days into pregnancy, with lower levels of POMC and GR promoter methylation. This was associated with increased GR mRNA expression. Other regions of the brain that also express POMC and GR, did not exhibit these epigenetic modifications. This study revealed that maternal undernutrition induces tissue specific epigenetic changes in fetal hypothalami which may contribute to disease in later life. Twins have been shown to have similar phenotypic characteristics as maternally undernourished offspring and therefore it has been suggested that they may also be programmed, but by intrauterine growth restriction. Consequently, extensive methylation and histone analysis of GR and POMC promoter regions was carried out in twin fetal hypothalami and compared to maternally undernourished groups. Interestingly, the decreased POMC and GR methylation of our amplicons in the maternally undernourished fetal hypothalami was also observed in twin fetal hypothalamic. This was concomitant with histone modifications and alterations in overall DNA methyltransferase activity. However, it was found that there were no changes in the POMC and GR mRNA expression levels in twin fetuses, but we postulate that this may occur later in life. To determine if changes in the fetal epigenetic status of hypothalamic GR and POMC impacted the adult progeny, tissues were obtained from adult offspring of maternally undernourished ewes. Epigenetic changes in the hypothalamic GR promoter observed in the fetal group persisted into adulthood, with concurrent increases in GR mRNA and GR protein expression. Of these groups the undernourished adult male offspring had decreased hypothalamic POMC expression and increased fat mass, changes that are consistent with an obese phenotype. The epigenetic and expression status of GR in the hippocampus and pituitary were modified, but in a tissue and sex specific manner. POMC epigenetic changes in the brain were complex, with various levels of epigenetic and expression changes. Overall periconceptional undernutrition induces hypothalamic specific changes in the epigenetic status of the GR gene which is known to regulate energy balance. Hypothalamic changes were persistent from the fetal stage into adulthood, with modifications in other tissues occurring after birth. These adaptations have the potential to increase the offspring’s propensity to develop obesity and altered stress regulation in later life.

Glucocorticoids are effective immunosuppressive and anti-inflammatory agents, but some aspects of stress appear to be proinflammatory. This study investigates this apparent paradox as it applies to stress exposure and the development of arthritis in a rat strain that has subnormal hypothalamic-pituitary-adrenal (HPA) responsiveness. Female Lewis rats were subjected to 1 week of rotating, psychological stressors for 5 h daily, beginning 7 days following inoculation with type [I collagen. The collagen-induced arthritis (CIA) group exposed lo stress showed reduced ankle width increase (p < 0.001) and decreased hindlimb severity scores (p < 0.001). At sacrifice, 2 days following stress termination, no differences in either measure remained and there was no difference in hind paw volume. However, the area of the tibia invaded by stroma, as quantitated by image analysis, was reduced in the stressed rats (p < 0.05). In animals exposed to stress, adrenal weights were increased (p < 0.005) and plasma corticosterone levels were elevated at sacrifice (p < 0.02). Both injected groups had significantly larger adrenal (p < 0.005) and lower thymus weights (p < 0.05) than did uninjected controls. Likewise, both CIA groups had reduced glucocorticoid receptor immunoreactivity in synovial membranes compared to controls (p < 0.001), suggesting that the Lewis rat's HPA deficiency may be intensified by glucocorticoid receptor downregulation during the induction of CIA. These data indicate that the responsiveness of the HPA axis to psychological stress in this strain is sufficient to alter disease progression.