A hypothalamus és az autonóm idegrendszer szabályozó mechanizmusaiban részt vevő agypályák topográfiája és neurokémiai karakterizálásuk = Topography and neurochemical characterization of hypothalamic and central autonomic pathways

1. Feltérképezték a hypothalamus orexin-termelő neuronjainak agytörzsi projekcióit, pályáit, valamint elsőként mutatták ki kapcsolatait az agytörzs valamennyi noradrenalint és adrenalint termelő sejtcsoportjának neuronjaival. A vizsgálat eredménye alapvető fontosságú neuromorfológiai adat a táplálékfelvétel, valamint az agyi catecholamine rendszer kapcsolatának tisztázására a szervezet energia-háztartásának szempontjából. 2. Különböző neuroanatómiai és immunhisztokémiai vizsgálatokkal először írták le a korábban ismeretlen mediális paralemniscális magot patkány, majom és az emberi alsó agytörzsben. Igazolták egy speciális peptiderg (TIP39-PTH2 receptor) rendszer jelenlétét e mag területén. Tisztázták a magcsoport afferens és efferens neuronális kapcsolatait és adatot szolgáltattak ezen mag neuronjainak az akusztikus stresszben vitt szerepéről. 3. Igazolták egy limbikus agykéregből leszálló, a gyomor működésében szerepet vivő pálya multiszinaptikus jelenlétét és topográfiáját: infralimbic anterior cingulate cortex - centralis amygdala - paraszimpatikus dorsalis vagus mag - n. vagus pálya. 4. Elsőként írták le a ""jóllakottság"" érzés (satiety) agypályáját a gyomor - n. vagus - nucleus tractus solitariii - hypothalamus nucleus dorsomedialis. Igazolták az ezen pályán belüli szignál transzdukcióban a glucagon-like peptide-1 és a prolactin-releasing hormon neuropeptidek meghatározó szerepét. | In order to attain major objectives of the project, a number of various neuromorphological techniques have been successfully applied to identify, localize and characterize neuronal pathway that interconnect the hypothalamus and lower brainstem nuclei. The major new findings and observations are the follows: 1) Pathways and lower brainstem projections of hypothalamic orexin-expressing neurons have been verified a mapped topographically. Termination and synaptic contacts of orexin-containing fibers have been demonstrated on adrenaline- and noradrenaline-expressing neurons in each lower brainstem catecholamine cell group indicating the existence of a descending hypothalamic pathway that influence the body energy metabolism by controlling the peripheral catecholamine outflow. 2) The afferent and efferent neuronal connections of the pontine medial paralemniscal nucleus have been first described in the rat, monkey and human brains providing evidence for the hypothalamic connections of this cell group that may participate in acoustic stress response. 3) The description of a multisynaptic pathway between the limbic system and the lower brainstem autonomic centers via hypothalamus and the amygdala has been completed in the present study demonstrating the functional importance of limbic cortical areas on the functional activity of the stomach. 4) The complete neuronal pathway of the “satiety signal” from the stomach to the hypothalamic regulatory center has been first verified by demonstrating of the ascending glucagon-like peptide-1 projections from the nucleus of the solitary tract to dorsomedial hypothalamic neurons. This may represent one of the most important link in the mechanism that control the central regulation of food intake

Colocalized neurotransmitters in the hindbrain cooperate in adaptation to chronic hypernatremia

Chronic hypernatremia activates the central osmoregulatory mechanisms and inhibits the function of the hypothalamic-pituitary-adrenal (HPA) axis. Noradrenaline (NE) release into the periventricular anteroventral third ventricle region (AV3V), the supraoptic (SON) and hypothalamic paraventricular nuclei (PVN) from efferents of the caudal ventrolateral (cVLM) and dorsomedial (cDMM) medulla has been shown to be essential for the hypernatremia-evoked responses and for the HPA response to acute restraint. Notably, the medullary NE cell groups highly coexpress prolactin-releasing peptide (PrRP) and nesfatin-1/NUCB2 (nesfatin), therefore, we assumed they contributed to the reactions to chronic hypernatremia. To investigate this, we compared two models: homozygous Brattleboro rats with hereditary diabetes insipidus (DI) and Wistar rats subjected to chronic high salt solution (HS) intake. HS rats had higher plasma osmolality than DI rats. PrRP and nesfatin mRNA levels were higher in both models, in both medullary regions compared to controls. Elevated basal tyrosine hydroxylase (TH) expression and impaired restraint-induced TH, PrRP and nesfatin expression elevations in the cVLM were, however, detected only in HS, but not in DI rats. Simultaneously, only HS rats exhibited classical signs of chronic stress and severely blunted hormonal reactions to acute restraint. Data suggest that HPA axis responsiveness to restraint depends on the type of hypernatremia, and on NE capacity in the cVLM. Additionally, NE and PrRP signalization primarily of medullary origin is increased in the SON, PVN and AV3V in HS rats. This suggests a cooperative action in the adaptation responses and designates the AV3V as a new site for PrRP's action in hypernatremia

Nesfatin-1/NUCB2 as a Potential New Element of Sleep Regulation in Rats.

STUDY OBJECTIVES: Millions suffer from sleep disorders that often accompany severe illnesses such as major depression; a leading psychiatric disorder characterized by appetite and rapid eye movement sleep (REMS) abnormalities. Melanin-concentrating hormone (MCH) and nesfatin-1/NUCB2 (nesfatin) are strongly co - expressed in the hypothalamus and are involved both in food intake regulation and depression. Since MCH was recognized earlier as a hypnogenic factor, we analyzed the potential role of nesfatin on vigilance. DESIGN: We subjected rats to a 72 h-long REMS deprivation using the classic flower pot method, followed by a 3 h-long 'rebound sleep'. Nesfatin mRNA and protein expressions as well as neuronal activity (Fos) were measured by quantitative in situ hybridization technique, ELISA and immunohistochemistry, respectively, in 'deprived' and 'rebound' groups, relative to controls sacrificed at the same time. We also analyzed electroencephalogram of rats treated by intracerebroventricularly administered nesfatin-1, or saline. RESULTS: REMS deprivation downregulated the expression of nesfatin (mRNA and protein), however, enhanced REMS during 'rebound' reversed this to control levels. Additionally, increased transcriptional activity (Fos) was demonstrated in nesfatin neurons during 'rebound'. Centrally administered nesfatin-1 at light on reduced REMS and intermediate stage of sleep, while increased passive wake for several hours and also caused a short-term increase in light slow wave sleep. CONCLUSIONS: The data designate nesfatin as a potential new factor in sleep regulation, which fact can also be relevant in the better understanding of the role of nesfatin in the pathomechanism of depression

Nesfatin-1/NUCB2 may participate in the activation of the hypothalamic-pituitary-adrenal axis in rats

Nesfatin-1 is an anorexigenic peptide originating from nucleobinding-2 (NUCB2) protein. Nesfatin-1/NUCB2-immunoreactive neurons are present in the hypothalamic paraventricular nucleus, the center of the stress-axis, and in the medullary A1 and A2 catecholamine cell groups. The A1 and A2 cell groups mediate viscerosensory stress information toward the hypothalamic paraventricular nucleus. They contain noradrenaline, but subsets of these neurons also express prolactin-releasing peptide acting synergistically with noradrenaline in the activation of the hypothalamic paraventricular nucleus during stress. We investigated the possible role of nesfatin-1/NUCB2 in the stress response. Intracerebro-ventricular administration of nesfatin-1 elevated both plasma adrenocorticotropin and corticosterone levels, while in vitro stimulation of the hypophysis was ineffective. Single, long-duration restraint stress activated (Fos positivity) many of the nesfatin-1/NUCB2-immunoreactive neurons in the parvocellular part of the hypothalamic paraventricular nucleus, evoked nesfatin-1/NUCB2 mRNA expression in the parvocellular part of the paraventricular nucleus and in the A1, but not in the A2 cell group. Nesfatin-1/NUCB2 was shown to co-localize in a high percentage of prolactin-releasing peptide producing neurons, in both medullary catecholamine cell groups further supporting its involvement in the stress response. Finally, bilateral adrenalectomy evoked an increasing nesfatin-1/NUCB2 mRNA expression, indicating that it is under the negative feedback of adrenal steroids. These data provide the first evidence for possible participation of nesfatin-1/NUCB2 in the stress-axis regulation, both at the level of the brainstem and in the hypothalamus

Hypothalamic Nesfatin-1 Resistance May Underlie the Development of Type 2 Diabetes Mellitus in Maternally Undernourished Non-obese Rats

Intrauterine growth retardation (IUGR) poses a high risk for developing late-onset, non-obese type 2 diabetes (T2DM). The exact mechanism underlying this phenomenon is unknown, although the contribution of the central nervous system is recognized. The main hypothalamic nuclei involved in the homeostatic regulation express nesfatin-1, an anorexigenic neuropeptide and identified regulator of blood glucose level. Using intrauterine protein restricted rat model (PR) of IUGR, we investigated, whether IUGR alters the function of nesfatin-1. We show that PR rats develop fat preference and impaired glucose homeostasis by adulthood, while the body composition and caloric intake of normal nourished (NN) and PR rats are similar. Plasma nesfatin-1 levels are unaffected by IUGR in both neonates and adults, but pro-nesfatin-1 mRNA expression is upregulated in the hypothalamus of adult PR animals. We find that centrally injected nesfatin-1 inhibits the fasting induced neuronal activation in the hypothalamic arcuate nucleus in adult NN rats. This effect of nesfatin-1 is not seen in PR rats. The anorexigenic effect of centrally injected nesfatin-1 is also reduced in adult PR rats. Moreover, chronic central nesfatin-1 administration improves the glucose tolerance and insulin sensitivity in NN rats but not in PR animals. Birth dating of nesfatin-1 cells by bromodeoxyuridine (BrDU) reveals that formation of nesfatin-1 cells in the hypothalamus of PR rats is disturbed. Our results suggest that adult PR rats acquire hypothalamic nesfatin-1-resistance, probably due to the altered development of the hypothalamic nesfatin-1 cells. Hypothalamic nesfatin-1-resistance, in turn, may contribute to the development of non-obese type T2DM

Topical analgesic, anti-inflammatory and antioxidant properties of Oxybaphus nyctagineus: Phytochemical characterization of active fractions

Ethnopharmacological relevance Oxybaphus nyctagineus (Michx.) Sweet has traditionally been used by several Native American tribes predominantly as a topical anti-inflammatory and analgesic agent. Aim of the study To evaluate the antioxidant, analgesic and anti-inflammatory activity of the extracts prepared from the aerial parts of Oxybaphus nyctagineus and to characterize the major chemical constituents of the bioactive extracts. Materials and methods Crude polar and apolar extracts (PCE and ACE) of the herb of Oxybaphus nyctagineus were prepared and tested in the models of the CFA-induced hyperalgesia in rat knee and carrageenan-induced paw edema in rat. To identify the active compounds, subfractions were prepared by column chromatography and subjected in vitro assays, such as antioxidant assays (DPPH, peroxynitrite (ONOO-) scavenging), and the LPS-induced IL-1β release test in human monocytes. Preparative HPLC was employed for the isolation of active substances, while phytochemical analysis was performed by mean of LC-MS/MS and NMR. Results The topically administered PCE and ACE of Oxybaphus nyctagineus demonstrated a significant analgesic and anti-inflammatory effect in the inflammation animal models. The subfraction A4 of ACE and the subfraction P5 of PCE considerably inhibited the LPS-induced IL-1β release in human monocytes, while the strongest activity was localized in the subfraction P5 in the antioxidant assays. The HPLC-MS/MS and NMR analysis revealed that 6-methoxyflavonol diglycosides, namely patuletin-3-O-robinobioside (1), 6-methoxykaempferol-3-O- robinobioside (2), spinacetin-3-O-robinobioside (3), and hydroxy-polyenoic fatty acids, namely corchorifatty acid B (4), 9-hydroxy-10E,12Z,15Z-octadecatrienoic acid (9-HOT acid) (5), and 9-hydroxy-10E,12Z-octadecadienoic acid (9-HOD acid) (6) were present in PCE, and in ACE as major compounds. Conclusion The results of this study established a pharmacological evidence for the traditional use of Oxybaphus nyctagineus as an anti-inflammatory agent used topically, and provided data on its phytochemical composition for the first time. © 2014 Elsevier Ireland Ltd. All rights reserved

Participation of melanin-concentrating hormone (MCH)-positive and MCH - negative nesfatin-1/NUCB2 (nesfatin) neurons in the sleep - wake cycle.

<p><b>A–C</b>. Illustrative pictures of the lateral hypothalamic area demonstrating the results of the triple fluorescent immunostainings for nesfatin (red), MCH (blue) and Fos (green) in a home cage kept (<b>A</b>), a rapid eye movement sleep (REMS) - deprived (<b>B</b>) and a REMS - deprived - sleep rebound (<b>C</b>) animal. Nesfatin/MCH double-positive neurons are pink (arrows), MCH - negative nesfatin neurons are red (arrowheads). Activated nesfatin/MCH neurons show white nuclei, activated nesfatin - positive, but MCH - negative neurons show yellow nuclei. Note that majority of the MCH - positive nesfatin neurons are activated (Fos - positive, arrows) by rebound, while only a few of the MCH - negative neurons showed Fos - positivity (arrowhead). Scale bar: 100 µm. <b>D</b>. Distribution of MCH - positive (N⁺/M⁺) and MCH - negative (N⁺/M^¯) neurons within the nesfatin producing cell population and percentage of activated (Fos - positive) cells after REMS deprivation followed by rebound. Data are shown as mean ± SEM, n = 5.</p

Effects of intracerebroventricularly administered nesfatin-1 on slow wave sleep and passive wake (PW) vigilance stages.

<p>The time spent and the number of episodes in light slow wave sleep (SWS1, <b>A</b> and <b>D</b>, respectively), deep slow wave sleep (SWS2, <b>B</b> and <b>E</b>, respectively) as well as in PW (<b>C</b> and <b>F</b>, respectively), per hour in the 2^nd–6^th hours of passive (light) phase. Data are presented as mean ± SEM, n = 6 per group, p*<0.05, p**<0.01.</p

Stress-related changes and energy balance of the experimental animals.

<p>Data show significantly increased levels of CRH mRNA in the hypothalamic paraventricular nucleus (<b>A</b>) and decreased body weights (<b>B</b>) both in the SP and LP kept animals, compared to HC, without difference between the above mentioned groups. <b>C</b>. Cumulative food intake shows no alterations. HC: home cage control, sacrificed with the animals kept on platforms, HCR: home cage control “rebound”, sacrificed at the same time as rebound groups, SP: small pot, SPR: small pot plus sleep rebound, LP: large pot, LPR: large pot plus sleep rebound groups. Data are shown as mean ± SEM, n = 10–11, p*<0.05, p**<0.01 <i>vs</i>. HC.</p