Important role of ventromedial hypothalamic carnitine palmitoyltransferase-1a in the control of food intake

Carnitine palmitoyltransferase-1 (CPT-1) liver isoform or CPT-1a is implicated in CNS control of food intake. However, the exact brain nucleus site(s) in mediating this action of CPT-1a has not been identified. In this report, we assess the role of CPT-1a in hypothalamic ventromedial nucleus (VMN). We stereotaxically injected an adenoviral vector containing CPT-1a coding sequence into the VMN of rats to induce overexpression and activation of CPT-1a. The VMN-selective activation of CPT-1a induced orexigenic effect, suggesting CPT-1a in the VMN is involved in the central control of feeding. Intracerebroventricular administration of etomoxir, a CPT-1 inhibitor, decreases food intake. Importantly, in the animals with VMN-overexpression of a CPT-1a mutant that antagonizes the CPT-1 inhibition by etomoxir, the anorectic response to etomoxir was attenuated. This suggests that VMN is involved in mediating the anorectic effect of central inhibition of CPT-1a. In contrast, Arc overexpression of the mutant did not alter etomoxir-induced inhibition of food intake, suggesting that Arc CPT-1a does not play significant roles in this anorectic action. Furthermore, in the VMN, CPT-1a appears to act downstream of hypothalamic malonyl-CoA action of feeding. Finally, we show that in the VMN, CPT-1 activity altered in concert with fasting and refeeding states, supporting a physiological role of CPT-1a in mediating the control of feeding. Taking together, CPT-1a in the hypothalamic VMN appears to play an important role in the central control of food intake. VMN-selective modulation of CPT-1a activity may therefore be a promising strategy in controlling food intake and maintaining normal body weight.

Agricultural land use and human presence around breeding sites increase stress-hormone levels and decrease body mass in barn owl nestlings.

Human activities can have a suite of positive and negative effects on animals and thus can affect various life history parameters. Human presence and agricultural practice can be perceived as stressors to which animals react with the secretion of glucocorticoids. The acute short-term secretion of glucocorticoids is considered beneficial and helps an animal to redirect energy and behaviour to cope with a critical situation. However, a long-term increase of glucocorticoids can impair e.g. growth and immune functions. We investigated how nestling barn owls (Tyto alba) are affected by the surrounding landscape and by human activities around their nest sites. We studied these effects on two response levels: (a) the physiological level of the hypothalamus-pituitary-adrenal axis, represented by baseline concentrations of corticosterone and the concentration attained by a standardized stressor; (b) fitness parameters: growth of the nestlings and breeding performance. Nestlings growing up in intensively cultivated areas showed increased baseline corticosterone levels late in the season and had an increased corticosterone release after a stressful event, while their body mass was decreased. Nestlings experiencing frequent anthropogenic disturbance had elevated baseline corticosterone levels, an increased corticosterone stress response and a lower body mass. Finally, breeding performance was better in structurally more diverse landscapes. In conclusion, anthropogenic disturbance affects offspring quality rather than quantity, whereas agricultural practices affect both life history traits.

The central histaminergic neurons in vitro, and their neuroprotective role in excitotoxic cell death in the postnatal rat hippocampus.

Histamine acts as a neurotransmitter in the central nervous system. Brain histamine in synthesized in neurons located to the posterior hypothalamus, from where these neurons send their projections to different parts of the brain. Released histamine participates in the regulation of several physiological functions such as arousal, attention and body homeostasis. Disturbances in the histaminergic system have been detected in diseases such as epilepsy, sleep disorders, anxiety, depression, Alzheimer’s disease, and schizophrenia. The purpose of this thesis was to develop optimal culture conditions for the histaminergic neurons, to study their detailed morphology, and to find out their significance in the kainic acid (KA)-induced neuronal death in the immature rat hippocampus. The morphology of the histaminergic neurons in vitro was comparable with the earlier findings. Histamine-containing vesicles were found in the axon but also in the cell body and dendrites suggesting a possibility for the somatodendritic release. Moreover, histamine was shown to be colocalized with the vesicular monoamine transporter 2 (VMAT2) suggesting that VMAT2 transports histamine to the subcellular storage vesicles. Furthermore, histamine was localized with γ-aminobutyric acid (GABA) in distinct storage vesicles and with neuropeptide galanin partly in the same storage vesicles suggesting different corelease mechanisms for GABA and galanin with histamine. In the organotypic hippocampal slice cultures, KA-induced neuronal death was first detected 12 h after the treatment being restricted mainly to the CA3 subregion. Moreover, cell death was irreversible, since the 48 h recovery period did not save the cells, but instead increased the damage. Finally, neuronal death was suggested to be necrotic, since intracellular apoptotic pathways were not activated, and the morphological changes detected with the electron microscopy were characteristic for necrosis. In the coculture system of the hippocampal and posterior hypothalamic slices, histaminergic neurons significantly decreased epileptiform burst activity and neuronal death in the hippocampal slices, this effect being mediated by histamine 1 (H1) and 3 (H3) receptors. In conclusion, the histaminergic neurons were maintained succesfully in the in vitro conditions exhibiting comparable morphological characteristics as detected earlier in vivo. Moreover, they developed functional innervations within the hippocampal slices in the coculture system. Finally, the KA-induced regionspecific, irreversible and necrotic hippocampal pyramidal cell damage was significantly decreased by the histaminergic neurons through H1 and H3 receptors.

Large-scale genomic analyses link reproductive aging to hypothalamic signaling, breast cancer susceptibility and BRCA1-mediated DNA repair.

Menopause timing has a substantial impact on infertility and risk of disease, including breast cancer, but the underlying mechanisms are poorly understood. We report a dual strategy in ∼70,000 women to identify common and low-frequency protein-coding variation associated with age at natural menopause (ANM). We identified 44 regions with common variants, including two regions harboring additional rare missense alleles of large effect. We found enrichment of signals in or near genes involved in delayed puberty, highlighting the first molecular links between the onset and end of reproductive lifespan. Pathway analyses identified major association with DNA damage response (DDR) genes, including the first common coding variant in BRCA1 associated with any complex trait. Mendelian randomization analyses supported a causal effect of later ANM on breast cancer risk (∼6% increase in risk per year; P = 3 × 10(-14)), likely mediated by prolonged sex hormone exposure rather than DDR mechanisms.

Le rôle de CRTC1 dans la prise de poids induite par les antipsychotiques

Les antipsychotiques atypiques, de deuxième génération, ont largement contribué à améliorer le traitement des patients souffrant de schizophrénie. Cependant, leur mécanisme d'action reste mal compris et leurs effets secondaires sont importants, notamment la prise de poids. CRTC1 (CREB-regulated transcription coactivator 1), aussi appelé TORC1 (transducers of regulated CREB activity 1), est un coactivateur de CREB. Il régule la transcription de Bdnf1 qui joue un rôle essentiel dans le contrôle de la balance énergétique dépendant du VMH 2, 3, 4. Nous pensons que CRTC1 est impliqué dans la prise de poids induite par certains antipsychotiques. En effet, il a été démontré que les souris Crtc1-/- devenaient hyperphagiques et obèses 5, 6, que la régulation de l'activité de CRTC1 se faisait par l'AMPK et que les antipsychotiques atypiques activaient cette kinase dans l'hypothalamus.7 L'AMPK de l'hypothalamus est liée à la régulation de la prise alimentaire, elle inverse l'action de la leptine, hormone anorexigène. Suite à ces constatations, nous proposons de suivre l'hypothèse de travail suivante : l'activation de l'AMPK par les antipsychotiques atypiques dans l'hypothalamus peut maintenir la phosphorylation de CRTC1 et le bloquer dans le cytoplasme, l'empêchant ainsi d'activer les gènes anorexigènes, comme Bdnf par exemple. En effet, la forme phosphorylée inactive de CRTC1 est séquestrée dans le cytoplasme et sa migration dans le noyau nécessite en même temps l'activation de la phosphatase calcineurine et l'inactivation des kinases de la famille de l'AMPK. Dans le travail que nous avons entamé, nous cherchons donc, par western blot et par immunohistochimie, à comprendre si les antipsychotiques atypiques inactivent CRTC1 en induisant sa phosphorylation par l'AMPK et sa rétention dans le cytoplasme.

Estudio del desarrollo y conectividad de las neuronas vasotocinérgicas en el prosencéfalo de pollo

En este Trabajo de Fin de Grado se estudia el origen embrionario de las distintas poblaciones neuronales que forman la amígdala medial extendida. La amígdala es una estructura del cerebro anterior involucrada en otorgar un significado emocional a los estímulos ambientales y en el control de distintos aspectos del comportamiento social (p.ej. comportamientos sexual, maternal, agresivo y afiliativo). Ante dichos estímulos, la amígdala pone en marcha una serie de reacciones de carácter motor, autonómico y endocrino que constituyen la respuesta emocional. Algunos desórdenes de carácter neuropsiquiátrico en humanos están relacionados con una disfunción en el control de las emociones y del comportamiento social, y varios de ellos se asocian a alteraciones en el desarrollo de la amígdala. El objetivo del presente trabajo ha sido investigar el origen de las neuronas de la amígdala medial extendida en embriones de pollo (E15 y E18) mediante ensayos de inmunocitoquímica, técnica utilizada para localizar las células que contienen el neuropéptido vasotocina (AVT) y proteínas reguladoras del desarrollo (la producida a partir del gen Otp) para ayudar en la delimitación de los distintos dominios embrionarios del prosencéfalo y distintas subdivisiones de la amígdala extendida. Los resultados de estos ensayos se combinaron con ensayos de trazado de conexiones para analizar la conectividad de las neuronas vasotocinérgicas de esta estructura. Los resultados obtenidos sugieren que las neuronas AVT-positivas podrían derivar del dominio Supra-Opto-Paraventricular (SPV), y algunas poblaciones alcanzarían su posición definitiva dentro del propio dominio por migración radial, mientras que otras invadirían otros dominios cerebrales por migración tangencial. En conclusión, la investigación proporciona importantes datos que clarifican aspectos relevantes del desarrollo y organización adulta de la amígdala extendida, y ayuda a establecer las bases para una mejor comprensión del control neural de las emociones y el comportamiento social en condiciones normales y patológicas.

Evidence for hypothalamic ketone body sensing: impact on food intake and peripheral metabolic responses in mice.

Monocarboxylates have been implicated in the control of energy homeostasis. Among them, the putative role of ketone bodies produced notably during high-fat diet (HFD) has not been thoroughly explored. In this study, we aimed to determine the impact of a specific rise in cerebral ketone bodies on food intake and energy homeostasis regulation. A carotid infusion of ketone bodies was performed on mice to stimulate sensitive brain areas for 6 or 12 h. At each time point, food intake and different markers of energy homeostasis were analyzed to reveal the consequences of cerebral increase in ketone body level detection. First, an increase in food intake appeared over a 12-h period of brain ketone body perfusion. This stimulated food intake was associated with an increased expression of the hypothalamic neuropeptides NPY and AgRP as well as phosphorylated AMPK and is due to ketone bodies sensed by the brain, as blood ketone body levels did not change at that time. In parallel, gluconeogenesis and insulin sensitivity were transiently altered. Indeed, a dysregulation of glucose production and insulin secretion was observed after 6 h of ketone body perfusion, which reversed to normal at 12 h of perfusion. Altogether, these results suggest that an increase in brain ketone body concentration leads to hyperphagia and a transient perturbation of peripheral metabolic homeostasis.

Sociodemographic, behavioral and genetic determinants of allostatic load in a Swiss population-based study.

Allostatic load (AL) is a marker of physiological dysregulation which reflects exposure to chronic stress. High AL has been related to poorer health outcomes including mortality. We examine here the association of socioeconomic and lifestyle factors with AL. Additionally, we investigate the extent to which AL is genetically determined. We included 803 participants (52% women, mean age 48±16years) from a population and family-based Swiss study. We computed an AL index aggregating 14 markers from cardiovascular, metabolic, lipidic, oxidative, hypothalamus-pituitary-adrenal and inflammatory homeostatic axes. Education and occupational position were used as indicators of socioeconomic status. Marital status, stress, alcohol intake, smoking, dietary patterns and physical activity were considered as lifestyle factors. Heritability of AL was estimated by maximum likelihood. Women with a low occupational position had higher AL (low vs. high OR=3.99, 95%CI [1.22;13.05]), while the opposite was observed for men (middle vs. high OR=0.48, 95%CI [0.23;0.99]). Education tended to be inversely associated with AL in both sexes(low vs. high OR=3.54, 95%CI [1.69;7.4]/OR=1.59, 95%CI [0.88;2.90] in women/men). Heavy drinking men as well as women abstaining from alcohol had higher AL than moderate drinkers. Physical activity was protective against AL while high salt intake was related to increased AL risk. The heritability of AL was estimated to be 29.5% ±7.9%. Our results suggest that generalized physiological dysregulation, as measured by AL, is determined by both environmental and genetic factors. The genetic contribution to AL remains modest when compared to the environmental component, which explains approximately 70% of the phenotypic variance.

Contribution of subcortical structures to cognition assessed with invasive electrophysiology in humans

Implantation of deep brain stimulation (DBS) electrodes via stereotactic neurosurgery has become a standard procedure for the treatment of Parkinson's disease. More recently, the range of neuropsychiatric conditions and the possible target structures suitable for DBS have greatly increased. The former include obsessive compulsive disease, depression, obesity, tremor, dystonia, Tourette's syndrome and cluster-headache. In this article we argue that several of the target structures for DBS (nucleus accumbens, posterior inferior hypothalamus, nucleus subthalamicus, nuclei in the thalamus, globus pallidus internus, nucleus pedunculopontinus) are located at strategic positions within brain circuits related to motivational behaviors, learning, and motor regulation. Recording from DBS electrodes either during the operation or post-operatively from externalized leads while the patient is performing cognitive tasks tapping the functions of the respective circuits provides a new window on the brain mechanisms underlying these functions. This is exemplified by a study of a patient suffering from obsessive-compulsive disease from whom we recorded in a flanker task designed to assess action monitoring processes while he received a DBS electrode in the right nucleus accumbens. Clear error-related modulations were obtained from the target structure, demonstrating a role of the nucleus accumbens in action monitoring. Based on recent conceptualizations of several different functional loops and on neuroimaging results we suggest further lines of research using this new window on brain functions.

Alpha- and gamma-melanocyte stimulating hormones in obesity. A study of the key central areas of metabolic regulation

The melanocortin system is an important regulator of feeding, energy metabolism,and cardiovascular function and it consists of the pro-opiomelanocortin (POMC) derived melanocyte stimulating hormones (α-, β- and γ-MSH) and their endogenous melanocortin receptors, MC1R to MC5R. In the hypothalamus, α-MSH reduces food intake, and increases energy expenditure and sympathetic tone by binding to MC4R. Mutations affecting the MC4R gene lead to obesity in mammals. On the other hand, the metabolic effects of MC3R stimulation using agonists such as the endogenously expressed γ-MSH have been less extensively explored. The main objective of this study was to investigate the long-term effects of increased melanocortin tone in key areas of metabolic regulation in the central nervous system (CNS) in order to investigate the sitespecific roles of both α-MSH and γ-MSH. The aim was to stereotaxically induce local overexpression of single melanocortin peptides using lentiviral vectors expressing α-MSH (LVi-α-MSH-EGFP) and γ-MSH (LVi-γ-MSH-EGFP). The lentiviral vectors were shown to produce a long-term overexpression and biologically active peptides in cell-based assays. The LVi-α-MSHEGFP was targeted to the arcuate nucleus in the hypothalamus of diet induced obese mice where it reduced weight gain and adiposity independently of food intake. When the nucleus tractus solitarus in the brainstem was targeted, the LVi-α-MSH-EGFP treatment was shown to cause a small decrease in adiposity, which did not impact weight development. However, the α-MSH treatment increased heart rate, which was attenuated by adrenergic receptor blockade indicative of increased sympathetic activity. The LVi-γ-MSH-EGFP was targeted to the hypothalamus where it decreased fat mass in mice eating the standard diet, but the effect was abated if animals consumed a high-fat Western type diet. When the diet induced obese mice were subjected again to the standard diet, the LVi-γ-MSH-EGFP treated animals displayed increased weight loss and reduced adiposity. These results indicate that the long-term central anti-obesity effects of α-MSH are independent of food intake. In addition, overexpression of α-MSH in the brain stem efficiently blocked the development of adiposity, but increased sympathetic tone. The evidence presented in this thesis also indicates that selective MC3R agonists such as γ-MSH could be potential therapeutics in combination with low fat diets.

Correlations between ANP concentrations in atria, plasma and cerebral structures and sodium chloride preference in Wistar rats

We determined whether ANP (atrial natriuretic peptide) concentrations, measured by radioimmunoassay, in the ANPergic cerebral regions involved in regulation of sodium intake and excretion and pituitary gland correlated with differences in sodium preference among 40 Wistar male rats (180-220 g). Sodium preference was measured as mean spontaneous ingestion of 1.5% NaCl solution during a test period of 12 days. The relevant tissues included the olfactory bulb (OB), the posterior and anterior lobes of the pituitary gland (PP and AP, respectively), the median eminence (ME), the medial basal hypothalamus (MBH), and the region anteroventral to the third ventricle (AV3V). We also measured ANP content in the right (RA) and left atrium (LA) and plasma. The concentrations of ANP in the OB and the AP were correlated with sodium ingestion during the preceding 24 h, since an increase of ANP in these structures was associated with a reduced ingestion and vice-versa (OB: r = -0.3649, P<0.05; AP: r = -0.3291, P<0.05). Moreover, the AP exhibited a correlation between ANP concentration and mean NaCl intake (r = -0.4165, P<0.05), but this was not the case for the OB (r = 0.2422). This suggests that differences in sodium preference among individual male rats can be related to variations of AP ANP level. Earlier studies indicated that the OB is involved in the control of NaCl ingestion. Our data suggest that the OB ANP level may play a role mainly in day-to-day variations of sodium ingestion in the individual rat

Faster gastric emptying of a liquid meal in rats after hypothalamic dorsomedial nucleus lesion

The effects of dorsomedial hypothalamic (DMH) nucleus lesion on body weight, plasma glucose levels, and the gastric emptying of a liquid meal were investigated in male Wistar rats (170-250 g). DMH lesions were produced stereotaxically by delivering a 2.0-mA current for 20 s through nichrome electrodes (0.3-mm tip exposure). In a second set of experiments, the DMH and the ventromedial hypothalamic (VMH) nucleus were lesioned with a 1.0-mA current for 10 s (0.1-mm tip exposure). The medial hypothalamus (MH) was also lesioned separately using a nichrome electrode (0.3-mm tip exposure) with a 2.0-mA current for 20 s. Gastric emptying was measured following the orogastric infusion of a liquid test meal consisting of physiological saline (0.9% NaCl, w/v) plus phenol red dye (6 mg/dl) as a marker. Plasma glucose levels were determined after an 18-h fast before the lesion and on the 7th and 15th postoperative day. Body weight was determined before lesioning and before sacrificing the rats. The DMH-lesioned rats showed a significantly faster (P<0.05) gastric emptying (24.7% gastric retention, N = 11) than control (33.0% gastric retention, N = 8) and sham-lesioned (33.5% gastric retention, N = 12) rats, with a transient hypoglycemia on the 7th postoperative day which returned to normal by the 15th postoperative day. In all cases, weight gain was slower among lesioned rats. Additional experiments using a smaller current to induce lesions confirmed that DMH-lesioned rats had a faster gastric emptying (25.1% gastric retention, N = 7) than control (33.4% gastric retention, N = 17) and VMH-lesioned (34.6% gastric retention, N = 7) rats. MH lesions resulted in an even slower gastric emptying (43.7% gastric retention, N = 7) than in the latter two groups. We conclude that although DMH lesions reduce weight gain, they do not produce consistent changes in plasma glucose levels. These lesions also promote faster gastric emptying of an inert liquid meal, thus suggesting a role for the DMH in the regulation of gastric motility

Neonatal handling induces anovulatory estrous cycles in rats

Since previous work has shown that stimulation early in life decreases sexual receptiveness as measured by the female lordosis quotient, we suggested that neonatal handling could affect the function of the hypothalamus-pituitary-gonadal axis. The effects of neonatal handling on the estrous cycle and ovulation were analyzed in adult rats. Two groups of animals were studied: intact (no manipulation, N = 10) and handled (N = 11). Pups were either handled daily for 1 min during the first 10 days of life or left undisturbed. At the age of 90 days, a vaginal smear was collected daily at 9:00 a.m. and analyzed for 29 days; at 9:00 a.m. on the day of estrus, animals were anesthetized with thiopental (40 mg/kg, ip), the ovaries were removed and the oviduct was dissected and squashed between 2 glass slides. The number of oocytes of both oviductal ampullae was counted under the microscope. The average numbers for each phase of the cycle (diestrus I, diestrus II, proestrus and estrus) during the period analyzed were compared between the two groups. There were no significant differences between intact and handled females during any of the phases. However, the number of handled females that showed anovulatory cycles (8 out of 11) was significantly higher than in the intact group (none out of 10). Neonatal stimulation may affect not only the hypothalamus-pituitary-adrenal axis, as previously demonstrated, but also the hypothalamus-pituitary-gonadal axis in female rats.

Role of sympathetic nervous system and neuropeptides in obesity hypertension

Obesity is the most common cause of human essential hypertension in most industrialized countries. Although the precise mechanisms of obesity hypertension are not fully understood, considerable evidence suggests that excess renal sodium reabsorption and a hypertensive shift of pressure natriuresis play a major role. Sympathetic activation appears to mediate at least part of the obesity-induced sodium retention and hypertension since adrenergic blockade or renal denervation markedly attenuates these changes. Recent observations suggest that leptin and its multiple interactions with neuropeptides in the hypothalamus may link excess weight gain with increased sympathetic activity. Leptin is produced mainly in adipocytes and is believed to regulate energy balance by acting on the hypothalamus to reduce food intake and to increase energy expenditure via sympathetic activation. Short-term administration of leptin into the cerebral ventricles increases renal sympathetic activity, and long-term leptin infusion at rates that mimic plasma concentrations found in obesity raises arterial pressure and heart rate via adrenergic activation in non-obese rodents. Transgenic mice overexpressing leptin also develop hypertension. Acute studies suggest that the renal sympathetic effects of leptin may depend on interactions with other neurochemical pathways in the hypothalamus, including the melanocortin-4 receptor (MC4-R). However, the role of this pathway in mediating the long-term effects of leptin on blood pressure is unclear. Also, it is uncertain whether there is resistance to the chronic renal sympathetic and blood pressure effects of leptin in obese subjects. In addition, leptin also has other cardiovascular and renal actions, such as stimulation of nitric oxide formation and improvement of insulin sensitivity, which may tend to reduce blood pressure in some conditions. Although the role of these mechanisms in human obesity has not been elucidated, this remains a fruitful area for further investigation, especially in view of the current "epidemic" of obesity in most industrialized countries.

The physiological role of AT1 receptors in the ventrolateral medulla

Neurons in the rostral and caudal parts of the ventrolateral medulla (VLM) play a pivotal role in the regulation of sympathetic vasomotor activity and blood pressure. Studies in several species, including humans, have shown that these regions contain a high density of AT1 receptors specifically associated with neurons that regulate the sympathetic vasomotor outflow, or the secretion of vasopressin from the hypothalamus. It is well established that specific activation of AT1 receptors by application of exogenous angiotensin II in the rostral and caudal VLM excites sympathoexcitatory and sympathoinhibitory neurons, respectively, but the physiological role of these receptors in the normal synaptic regulation of VLM neurons is not known. In this paper we review studies which have defined the effects of specific activation or blockade of these receptors on cardiovascular function, and discuss what these findings tell us with regard to the physiological role of AT1 receptors in the VLM in the tonic and phasic regulation of sympathetic vasomotor activity and blood pressure.