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Background: The guidelines of brain death determination vary across countries. Our aim was to compare diagnostic procedures of brain death determination in adults among five countries., Method: Consecutive comatose patients who received brain death determination from June 2018 to June 2020 were included. The technical specifications, completion rates and positive rates of brain death determination according to criteria of different countries were compared. The accuracy, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of each ancillary test for the identification of brain death diagnosed according to different criteria were investigated., Results: One hundred and ninety nine patients were included in this study. One hundred and thirty one (65.8%) patients were diagnosed with brain death according to French criteria, 132 (66.3%) according to Chinese criteria, and 135 (67.7%) according to criteria of USA, UK and Germany. The sensitivity and PPV of electroencephalogram (92.2% - 92.3%) and somatosensory evoked potential (95.5% - 98.5%) were higher than transcranial Doppler (84.3% - 86.0%)., Conclusions: The criteria of brain death in China and France are comparatively stricter than in USA, UK and Germany. The discrepancy in brain death determination between clinical assessments and additional confirmation of ancillary tests is small.

Background: Alpha-synuclein (α-syn) is involved in pathology of Parkinson's disease, and 90% of α-syn in Lewy bodies is phosphorylated at serine 129 (pS129 α-syn)., Objective: To assess behavior impairments and brain levels of α-syn and pS129 α-syn in mice overexpressing human α-syn under Thy1 promoter (Thy1-α-syn) and wild type (wt) littermates., Methods: Motor and non-motor behaviors were monitored, brain human α-syn levels measured by ELISA, and α-syn and pS129 α-syn mapped by immunohistochemistry., Results: Male and female wt littermates did not show differences in the behavioral tests. Male Thy1-α-syn mice displayed more severe impairments than female counterparts in cotton nesting, pole tests, adhesive removal, finding buried food, and marble burying. Concentrations of human α-syn in the olfactory regions, cortex, nigrostriatal system, and dorsal medulla were significantly increased in Thy1-α-syn mice, higher in males than females. Immunoreactivity of α-syn was not simply increased in Thy1-α-syn mice but had altered localization in somas and fibers in a few brain areas. Abundant pS129 α-syn existed in many brain areas of Thy1-α-syn mice, while there was none or only a small amount in a few brain regions of wt mice. The substantia nigra, olfactory regions, amygdala, lateral parabrachial nucleus, and dorsal vagal complex displayed different distribution patterns between wt and transgenic mice, but not between sexes., Conclusion: The severer abnormal behaviors in male than female Thy1-α-syn mice may be related to higher brain levels of human α-syn, in the absence of sex differences in the altered brain immunoreactivity patterns of α-syn and pS129 α-syn.

Extensive use of triazophos for the chemical control of pests in agriculture or aquaculture might strongly disturb the aquatic environment due to residue accumulation through various routes like surface run-off, spray-drift and effluent from factories, which have potential negative effects to non-target aquatic organisms. Previous studies have documented the antioxidative effects of triazophos to mammals, however, the oxidative toxicity of triazophos to fish has not been adequately studied to date. Thus, an acute exposure (96 h) to triazophos at different concentrations of 0.06, 0.3 and 1.5 mg/L (corresponding to 1/50th, 1/10th and 1/2th of 96 h-LC 50 , respectively), was conducted to investigate the triazophos-induced oxidative stress in adult zebrafish (Danio rerio). The results showed that the time- and dose-dependent induction of oxidative stress except for the ROS level in liver after 24 and 48 h exposure, as indicated by increased reactive oxygen species (ROS), malondialdehyde (MDA) level and a compromised antioxidant defense system, including increased superoxide dismutase (SOD) activity, catalase (CAT) activity, glutathione (GSH) content as well as the increased at first and decreased afterwards genes expression (Sod1, Sod2, Cat and Gpx) in brain. Simultaneously, ROS and MDA showed an increased trend, SOD activity, CAT activity and GSH content showed a trend of increasing at 24 h and decreasing at 48 h, and then increasing at 96 h and Sod1, Sod2, Cat and Gpx gene showed decreasing at first and then increasing in liver tissue. The present study concluded that the damage of the antioxidant system by triazophos induced oxidative stress in the brain and liver of zebrafish with concomitant lipid peroxidation, which is an important mechanism underlying the triazophos-induced acute toxicity., (Copyright © 2019 Elsevier Inc. All rights reserved.)

High-protein diet (HPD) curtails obesity and/or fat mass, but it is unknown whether it reverses neuroinflammation or alters glucose levels, CCK sensitivity, and gut microbiome in rats fed a Western diet (WD)-induced obesity (DIO). Male rats fed a WD (high fat and sugar) for 12 wk were switched to a HPD for 6 wk. Body composition, food intake, meal pattern, sensitivity to intraperitoneal CCK-8S, blood glucose, brain signaling, and cecal microbiota were assessed. When compared with a normal diet, WD increased body weight (9.3%) and fat mass (73.4%). CCK-8S (1.8 or 5.2 nmol/kg) did not alter food intake and meal pattern in DIO rats. Switching to a HPD for 6 wk reduced fat mass (15.7%) with a nonsignificantly reduced body weight gain, normalized blood glucose, and decreased feeding after CCK-8S. DIO rats on the WD or switched to a HPD showed comparable microbial diversity. However, in HPD versus WD rats, there was enrichment of 114 operational taxonomic units (OTUs) and depletion of 188 OTUs. Of those, Akkermansia muciniphila (enriched on a HPD), an unclassified Clostridiales, a member of the RF39 order, and a Phascolarctobacterium were significantly associated with fat mass. The WD increased cytokine expression in the hypothalamus and dorsal medulla that was unchanged by switching to HPD. These data indicate that HPD reduces body fat and restores glucose homeostasis and CCK sensitivity, while not modifying brain inflammation. In addition, expansion of cecal Akkermansia muciniphila correlated to fat mass loss may represent a potential peripheral mechanism of HPD beneficial effects.

The subthalamic nucleus regulates motor and neurocognitive functions. Because of its small size and close proximity to other small subcortical structures, it has been a challenge to localize and visualize it using MRI. Here, we sought to define the optimal MRI scan method and visualization plane for locating the subthalamic nucleus on MRI images and to further delineate the geometric dimensions of the subthalamic nucleus and their correlation with age, laterality, and sex. Healthy volunteers received axial, sagittal, and coronal T2_3D_DRIVE CLEAR, coronal T1-WI, coronal T2FLAIR, coronal T2, and coronal SWI sequence. The coronal T2-3D-DRIVE CLEAR images were compared with the Schaltenbrand-Wahren Atlas for Stereotaxy of the Human Brain for localizing the subthalamic nucleus. The best visualization plane with the largest sectional area and the most distinct outline was obtained and region of interest was delineated manually on the basis of the contours of the bilateral subthalamic nuclei in T2-WI images. T2-3D-DRIVE CLEAR in the coronal view showed optimal visualization of the subthalamic nucleus and indicated that the subthalamic nucleus showed three morphological types: the double convex lens type (172, 64%), the ram's horn type (62, 23%), and the willow leaf type (34, 13%). There were no statistically significant differences because of laterality, sex, and age in the sectional area, and maximal long and short diameter of the subthalamic nucleus. On the basis of our results, the current study has shown that T2-3D-DRIVE CLEAR in the coronal view provides optimal visualization of the subthalamic nucleus, which shows three distinct morphological types on MRI images, and there is no statistically significant difference in the geometric dimensions of the subthalamic nucleus because of laterality, sex, and age in normal individuals.

Abdominal surgery inhibits food intake and induces c-Fos expression in the hypothalamic and medullary nuclei in rats. Rikkunshito (RKT), a Kampo medicine improves anorexia. We assessed the alterations in meal microstructure and c-Fos expression in brain nuclei induced by abdominal surgery and the modulation by RKT in mice. RKT or vehicle was gavaged daily for 1 week. On day 8 mice had no access to food for 6-7 h and were treated twice with RKT or vehicle. Abdominal surgery (laparotomy-cecum palpation) was performed 1-2 h before the dark phase. The food intake and meal structures were monitored using an automated monitoring system for mice. Brain sections were processed for c-Fos immunoreactivity (ir) 2-h after abdominal surgery. Abdominal surgery significantly reduced bouts, meal frequency, size and duration, and time spent on meals, and increased inter-meal interval and satiety ratio resulting in 92-86% suppression of food intake at 2-24 h post-surgery compared with control group (no surgery). RKT significantly increased bouts, meal duration and the cumulative 12-h food intake by 11%. Abdominal surgery increased c-Fos in the prelimbic, cingulate and insular cortexes, and autonomic nuclei, such as the bed nucleus of the stria terminalis, central amygdala, hypothalamic supraoptic (SON), paraventricular and arcuate nuclei, Edinger-Westphal nucleus (E-W), lateral periaqueduct gray (PAG), lateral parabrachial nucleus, locus coeruleus, ventrolateral medulla and nucleus tractus solitarius (NTS). RKT induced a small increase in c-Fos-ir neurons in the SON and E-W of control mice, and in mice with surgery there was an increase in the lateral PAG and a decrease in the NTS. These findings indicate that abdominal surgery inhibits food intake by increasing both satiation (meal duration) and satiety (meal interval) and activates brain circuits involved in pain, feeding behavior and stress that may underlie the alterations of meal pattern and food intake inhibition. RKT improves food consumption post-surgically that may involve modulation of pain pathway.

The autonomic nervous system (ANS) conveys neuronal input from the brain to the stomach. We investigated mechanisms through which urocortin 1 (UCN1) injected intracerebroventricularly (ICV, 300 pmol/rat) inhibits circulating ghrelin in rats. This was achieved by assessing (1) the induction of c-fos gene expression as a marker of neuronal activation in specific hypothalamic and caudal brainstem regulating ANS; (2) the influence of vagotomy and pharmacological blockade of central and peripheral α- and β-adrenergic receptor (AR) on ICV UCN1-induced reduction of plasma ghrelin levels (determined by ELISA); and (3) the relevance of this pathway in the feeding response to a fast in rats. UCN1 increased c-fos mRNA expression in key brain sites influencing sympathetic activity namely the hypothalamic paraventricular and ventromedial nuclei, locus coeruleus, nucleus of the solitary tract, and rostral ventrolateral medulla, by 16-, 29-, 6-, 37-, and 13-fold, respectively. In contrast, the dorsal motor nucleus of the vagus had little c-fos mRNA expression and ICV UCN1 induced a similar reduction in acylated ghrelin in the sham-operated (31%) and vagotomized (41%) rats. An intraperitoneal (IP) injection of either a non-selective α- or selective α2-AR antagonist reduced, while a selective α2-AR agonist enhanced ICV UCN1-induced suppression of plasma acylated ghrelin levels. In addition, IP injection of a non-selective β- or selective β1-AR agonist blocked, and selective β1-AR antagonist augmented, the ghrelin response to ICV UCN1. The IP injections of a selective α1- or non-selective β or β2-AR antagonists, or any of the pretreatments given ICV had no effect. ICV UCN1 reduced the 2-h food intake in response to a fast by 80%, and this effect was partially prevented by a selective α2-AR antagonist. These data suggest that ICV UCN1 reduces plasma ghrelin mainly through the brain sympathetic component of the ANS and peripheral AR specifically α2-AR activation and inactivation of β1-AR. The α2-AR pathway contributes to the associated reduction in food intake., (Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Intracerebroventricular injection of stable somatostatin (SST) agonists stimulates food and water intake in rats. We investigated the receptor subtype(s) involved in the dipsogenic effect of intracerebroventricular injection of SST agonists, mechanisms of action, and role. In nonfasted and non-water-deprived male rats with chronic intracerebroventricular cannula, intake of water without food or food without water was monitored separately to avoid any interactions compared with intracerebroventricular vehicle. SST-14 and cortistatin (CST-14) (1 μg/rat icv) increased water intake by 3.1- and 2.7-fold, respectively, while both peptides did not alter food intake at 1 h postinjection in the light phase. By contrast, the stable pan-somatostatin agonist ODT8-SST (1 μg/rat icv) increased both water and food intake by 4.9- and 3.7-fold, respectively. S-346-011, a selective receptor 2 (sst2) agonist (1 μg/rat icv) induced water ingestion, while sst1 or sst4 agonist, injected under the same conditions, did not. The sst2 antagonist S-406-028 (1 μg/rat icv) prevented the 1-h water intake induced by intracerebroventricular ODT8-SST and CST-14. Losartan (100 μg/rat icv), an angiotensin receptor 1 (AT1) antagonist, completely blocked the water consumption induced by intracerebroventricular ODT8-SST, whereas intracerebroventricular injection of S-406-028 did not modify the intracerebroventricular ANG II-induced dipsogenic response. The sst2 antagonist reduced by 40% the increase of the 3-h water intake in the early dark phase. These data indicate that SST-14 and CST-14 interact with sst2 to exert a potent dipsogenic effect, which is mediated downstream by angiotensin-AT1 signaling. These data also indicate that sst2 activation by brain SST-14 and/or CST-14 may play an important role in the regulation of drinking behavior.

Tail pinch stimulates food intake in rats. We investigated brain mechanisms of this response and the influence of repeated exposure. Sprague-Dawley rats received acute (5 min) or repeated (5 min/day for 14 days) tail pinch using a padded clip. Acute tail pinch increased 5-min food intake compared with control (0.92 ± 0.2 vs. 0.03 ± 0.01 g, P < 0.01). This response was inhibited by 76% by intracerebroventricular injection of BIBP-3226, a neuropeptide Y1 (NPY1) receptor antagonist, increased by 48% by astressin-B, a corticotropin-releasing factor (CRF) receptor antagonist, and not modified by S-406-028, a somatostatin subtype 2 antagonist. After the 5-min tail pinch without food, blood glucose rose by 21% (P < 0.01) while changes in plasma acyl ghrelin (+41%) and adrenocorticotropic hormone (+37%) were not significant. Two tail pinches (45 min apart) activate pontine and hindbrain catecholaminergic and hypothalamic paraventricular CRF neurons. After 14 days of repeated tail pinch, the 5-min orexigenic response was not significantly different from days 2 to 11 but reduced by 50% thereafter (P < 0.001). Simultaneously, the 5-min fecal pellet output increased during the last 5 days compared with the first 5 days (+58%, P < 0.05). At day 14, the body weight gain was reduced by 22%, with a 99% inhibition of fat gain and a 25% reduction in lean mass (P < 0.05). The orexigenic response to acute 5-min tail pinch is likely to involve the activation of brain NPY1 signaling, whereas that of CRF tends to dampen the acute response and may contribute to increased defecation and decreased body weight gain induced by repeated tail pinch.

Mice and rats are widely used in stress-related behavioral studies while little is known about the distribution of the stress hormone, corticotropin-releasing factor (CRF) in the mouse brain. We developed and characterized a novel rat/mouse CRF polyclonal antibody (CURE ab 200101) that was used to detect and compare the brain distributions of CRF immunoreactivity in naïve and colchicine-treated rats and mice. We also assessed whether the visceral stressor of abdominal surgery activated brain CRF neurons using double labeling of Fos/CRF in naïve rats. CRF-ir neurons were visualized in the cortex, bed nucleus of the stria terminalis, central amygdala, hypothalamic paraventricular nucleus (PVN), Barrington's nucleus and dorsolateral tegmental area in naïve rats. CRF-immunoreactive (ir) neurons in the mouse brain were detected only after colchicine. The pattern shows fundamental similarity compared to the colchicine-treated rat brain, however, there were differences with a lesser distribution in both areas and density except in the lateral septum and external subnucleus of the lateral parabrachial nucleus which contained more CRF-ir neurons in mice, and CRF-ir neurons in the dorsal motor nucleus of the vagus were found only in mice. Abdominal surgery in naïve rats induced Fos-ir in 30% of total CRF-ir neurons in the PVN compared with control (anesthesia alone) while Fos was not co-localized with CRF in other brain nuclei. These data indicate that CRF-ir distribution in the brain displays similarity as well as distinct features in mice compared to rats that may underlie some differential stress responses. Abdominal surgery activates CRF-ir neurons selectively in the PVN of rats without colchicine treatment., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Nesfatin-1 reduces food intake when injected centrally in rodents. We recently described wide distribution of nucleobindin2 (NUCB2)/nesfatin-1 immunoreactivity in rat brain autonomic nuclei activated by various stressors. We used C57BL/6 mice to localize brain NUCB2/nesfatin-1 immunoreactivity and assessed activation of NUCB2/nesfatin 1 neurons after water avoidance stress (WAS). Gastric emptying of a non-nutrient liquid was also determined. NUCB2/nesfatin-1 immunoreactivity was detected in cortical areas including piriform, insular, cingulate and somatomotor cortices, the limbic system including amygdaloid nuclei, hippocampus and septum, the basal ganglia, bed nucleus of the stria terminalis, the thalamus including paraventricular and parafascicular nuclei, the hypothalamus including supraoptic, periventricular, paraventricular (PVN), arcuate nuclei and ventromedial and lateral hypothalamic areas. Intensely labeled NUCB2/nesfatin-1 neurons were detected in a previously undefined region which we named intermediate dorsomedial hypothalamus. In the brainstem, NUCB2/nesfatin-1 immunoreactivity was detected in the raphe nuclei, Edinger-Westphal nucleus, locus coeruleus (LC), lateral parabrachial nucleus, ventrolateral medulla (VLM) and dorsal vagal complex. WAS induced Fos expression in 35% of NUCB2/nesfatin-1-immunoreactive neurons in the PVN, 50% in the LC, 54% in the rostral raphe pallidus, 58% in the VLM, 39% in the middle part of the nucleus of the solitary tract (NTS) and 33% in the caudal NTS. Nesfatin-1 injected intracerebroventricularly significantly decreased gastric emptying. These data showed that NUCB2/nesfatin-1 immunoreactivity is distributed in mouse brain areas involved in the regulation of stress response and visceral functions activated by an acute psychological stressor suggesting that nesfatin-1 might play a role in the efferent component of the stress response., (Published by Elsevier B.V.)

Kisspeptin is distributed not only in brain areas for regulating reproduction but also in nuclei involved in feeding control. Whether kisspeptin alters food intake is unknown in mice. We examined how kisspeptin-10 influences feeding after intracerebroventricular injection in mice using automated monitoring. Kisspeptin-10 (0.3, 1, and 3 μg/mouse) dose-dependently inhibited the feeding response to an overnight fast by 50, 95, and 90% respectively, during the 2-3 h period postinjection. The 1μg/mouse dose reduced the 4-h cumulative food intake by 28% whereas intraperitoneal injection (10 μg/mouse) did not. The decreased 4-h food intake was due to reduced meal frequency (-45%/4 h), whereas meal size and gastric emptying were not altered. These data suggest that kisspeptin may be a negative central regulator of feeding by increasing satiety.

We recently reported that the oligosomatostatin receptor agonist, ODT8-SST increases food intake in rats via the somatostatin 2 receptor (sst(2)). We characterized ingestive behavior following intracerebroventricular (icv) injection of a selective sst(2) agonist in freely fed mice during the light phase. The sst(2) agonist (0.01, 0.03, 0.1, 0.3 or 1μg/mouse) injected icv under short inhalation anesthesia dose-dependently increased cumulative light phase food intake over 4h compared to vehicle with a 3.1-times increase at 1μg/mouse (p<0.05). Likewise, the sst(2,3,5) agonist octreotide (0.3 or 1μg/mouse) dose-dependently increased 4-h food intake, whereas selective sst(1) or sst(4) agonists at 1μg/mouse did not. In vehicle-treated mice, high fat diet increased caloric intake/4h by 2.8-times compared to regular diet (p<0.05) and values were further increased 1.4-times/4h by the sst(2) agonist. Automated continuous assessment of food intake established a 6.6-times higher food intake during the dark phase due to increased number of meals, meal size, meal duration and rate of ingestion compared to non-treated mice during the light phase. During the first 4h post icv sst(2) agonist injection, mice had a 57% increase in number of meals with a 60% higher rate of ingestion, and a 61% reduction in inter-meal intervals, whereas meal sizes were not altered compared to vehicle. These data indicate that the activation of brain sst(2) receptors potently stimulates the light phase ingestive behavior under basal or high fat diet-stimulated conditions in mice. The shortened inter-meal interval suggests an inhibitory effect of the sst(2) agonist on "satiety", whereas "satiation" is not altered as indicated by normal meal size., (Published by Elsevier Inc.)

Central activation of somatostatin (sst) receptors by oligosomatostatin analogs inhibits growth hormone and stress-related rise in catecholamine plasma levels while stimulating grooming, feeding behaviors, gastric transit and acid secretion, which can be mimicked by selective sst(2) receptor agonist. To evaluate the pattern of neuronal activation induced by peptide sst receptor agonists, we assessed Fos-expression in rat brain after intracerebroventricular (i.c.v.) injection of a newly developed selective sst(2) agonist compared to the oligosomatostatin ODT8-SST, a pan-sst(1-5) agonist. Ninety min after injection of vehicle (10 microl) or previously established maximal orexigenic dose of peptides (1 microg=1 nmol/rat), brains were assessed for Fos-immunohistochemistry and doublelabeling. Food and water were removed after injection. The sst(2) agonist and ODT8-SST induced a similar Fos distribution pattern except in the arcuate nucleus where only the sst(2) agonist increased Fos. Compared to ODT8-SST, the sst(2) agonist induced higher Fos-expression by 3.7-times in the basolateral amygdaloid nucleus, 1.2-times in the supraoptic nucleus (SON), 1.6-times in the magnocellular paraventricular hypothalamic nucleus (mPVN), 4.1-times in the external lateral parabrachial nucleus, and 2.6-times in both the inferior olivary nucleus and superficial layer of the caudal spinal trigeminal nucleus. Doublelabeling in the hypothalamus showed that ODT8-SST activates 36% of oxytocin, 63% of vasopressin and 79% of sst(2) immunoreactive neurons in the mPVN and 28%, 55% and 25% in the SON, respectively. Selective activation of sst(2) receptor results in a more robust neuronal activation than the pan-sst(1-5) agonist in various brain regions that may have relevance in sst(2) mediated alterations of behavioral, autonomic and endocrine functions., (Published by Elsevier B.V.)

Chronic stress and persistently high glucocorticoid levels can induce brain atrophy. Corticotropin-releasing factor (CRF)-overexpressing (OE) mice are a genetic model of chronic stress with elevated brain CRF and plasma corticosterone levels and Cushing's syndrome. The brain structural alterations in the CRF-OE mice, however, are not well known. We found that adult male and female CRF-OE mice had significantly lower whole brain and cerebellum weights than their wild type (WT) littermates (347.7+/-3.6mg vs. 460.1+/-4.3mg and 36.3+/-0.8mg vs. 50.0+/-1.3mg, respectively) without sex-related difference. The epididymal/parametrial fat mass was significantly higher in CRF-OE mice. The brain weight was inversely correlated to epididymal/parametrial fat weight, but not to body weight. Computerized image analysis system in Nissl-stained brain sections of female mice showed that the anterior cingulate and sensorimotor cortexes of CRF-OE mice were significantly thinner, and the volumes of the hippocampus, hypothalamic paraventricular nucleus and amygdala were significantly reduced compared to WT, while the locus coeruleus showed a non-significant increase. Motor functions determined by beam crossing and gait analysis showed that CRF-OE mice took longer time and more steps to traverse a beam with more errors, and displayed reduced stride length compared to their WT littermates. These data show that CRF-OE mice display brain size reduction associated with alterations of motor coordination and an increase in visceral fat mass providing a novel animal model to study mechanisms involved in brain atrophy under conditions of sustained elevation of brain CRF and circulating glucocorticoid levels., (Copyright 2010 Elsevier Ireland Ltd. All rights reserved.)

Abdominal surgery-induced postoperative gastric ileus is well established to induce Fos expression in specific brain nuclei in rats within 2-h after surgery. However, the phenotype of activated neurons has not been thoroughly characterized. Nesfatin-1 was recently discovered in the rat hypothalamus as a new anorexigenic peptide that also inhibits gastric emptying and is widely distributed in rat brain autonomic nuclei suggesting an involvement in stress responses. Therefore, we investigated whether abdominal surgery activates nesfatin-1-immunoreactive (ir) neurons in the rat brain. Two hours after abdominal surgery with cecal palpation under short isoflurane anesthesia or anesthesia alone, rats were transcardially perfused and brains processed for double immunohistochemical labeling of Fos and nesfatin-1. Abdominal surgery, compared to anesthesia alone, induced Fos expression in neurons of the supraoptic nucleus (SON), paraventricular nucleus (PVN), locus coeruleus (LC), Edinger-Westphal nucleus (EW), rostral raphe pallidus (rRPa), nucleus of the solitary tract (NTS) and ventrolateral medulla (VLM). Double Fos/nesfatin-1 labeling showed that of the activated cells, 99% were nesfatin-1-immunoreactive in the SON, 91% in the LC, 82% in the rRPa, 74% in the EW and VLM, 71% in the anterior parvicellular PVN, 47% in the lateral magnocellular PVN, 41% in the medial magnocellular PVN, 14% in the NTS and 9% in the medial parvicellular PVN. These data established nesfatin-1 immunoreactive neurons in specific nuclei of the hypothalamus and brainstem as part of the neuronal response to abdominal surgery and suggest a possible implication of nesfatin-1 in the alterations of food intake and gastric transit associated with such a stressor., ((c) 2009 Elsevier Inc. All rights reserved.)

Nesfatin-1 is one of the peptide products of posttranslational processing of the nucleobindin-2 (NUCB2) gene, suggested to have physiological relevance to suppress food intake and body weight gain in rats. Nesfatin-1-immunoreactive cells have been found in distinct nuclei in the rat brain related to circuitries regulating food intake. Here, we report novel yet undescribed localization of NUCB2/nesfatin-1 at the mRNA and protein level in the rat central nervous system. Immunohistochemical staining revealed the localization of NUCB2/nesfatin-1 in the piriform and insular cortex, endopiriform nucleus, nucleus accumbens, lateral septum, bed nucleus of stria terminalis, central amygdaloid nucleus, medial preoptic area, dorsal raphe nucleus, ambiguus nucleus, ventrolateral medulla and gigantocellular reticular nucleus, as well as Purkinje-cells of the cerebellum. In the spinal cord, nesfatin-1 immunoreactivity (IR) was found in both sympathetic and parasympathetic preganglionic neuronal groups and in the dorsal area X from lower thoracic to sacral segments. The immunohistochemical results were confirmed by RT-PCR in the central amygdaloid nucleus, nucleus accumbens, cerebellum and lumbar spinal cord microdissected by punch technique. The features and distributions of nesfatin-1 IR and mRNA expression in the brain and spinal cord suggest that NUCB2/nesfatin-1 could play a wider role in autonomic regulation of visceral-endocrine functions besides food intake.

Little is known about the chemical coding of the brain neuronal circuitry activated by nociceptive signals of visceral origin. We characterized brain nuclei activated during isovolumetric phasic distension of the proximal colon (10 ml, 30 s on/off for 10 min) in conscious male rats, using Fos as a marker of neuronal activation and dual immunohistochemistry to visualize co-localization of Fos expression and oxytocin (OT), arginine-vasopressin (AVP), corticotrophin-releasing factor (CRF) or tyrosine hydroxylase (TH). Proximal colon distension, compared with sham distension, induced a robust increase in Fos-like immunoreactive (IR) neurons in the paraventricular nucleus (PVN), supraoptic nucleus (SON) and accessory neurosecretory nuclei of the hypothalamus, nucleus of the solitary tract (NTS) and ventrolateral medulla (VLM), and to a lower extent, in the locus coeruleus (LC) and Barrington nucleus. Fos-IR neurons in the PVN after colon distension were identified in 81% of OT-IR, 18% AVP-IR and 16% CRF-IR neurons, while in the SON it represented 36% of OT-IR and 16% AVP-IR. Catecholaminergic cell groups in the pons (LC) and medulla (VLM, NTS) were also activated by proximal colon distension. Of the TH-IR neurons in VLM and NTS, 74% and 42% respectively were double labeled. These results indicate that colon distension stimulates OT-, AVP- and CRF-containing hypothalamic neurons, likely involved in the integration of colonic sensory information to modulate autonomic outflow and pain-related responses. Activation of medullary catecholaminergic centers might reflect the afferent and efferent limbs of the functional responses associated to visceral pain.

We assessed brain nuclei activated during noxious mechanical distension of the proximal colon in conscious rats, using Fos as a marker of neuronal activation, and functional reflex changes in gastric emptying associated to colon distension. The role of capsaicin-sensitive afferents in Fos and gastric responses to distension was also investigated. Compared with sham distension, isovolumetric phasic distension of the proximal colon (10 ml, 30 s on/off for 10 min) increased significantly Fos expression 1 h after distension in selective brain areas, most prominently, the paraventricular and supraoptic nuclei of the hypothalamus (13-fold and 80-fold, respectively), the locus coeruleus-Barrington's nucleus complex (2-fold), area postrema (7-fold) and the nucleus tractus solitarius (4-fold). Increased Fos expression was also observed in the cingulate cortex, posterior paraventricular nucleus of the thalamus, periaqueductal gray and ventrolateral medulla. Distension of the proximal colon significantly inhibited gastric emptying by 82% and 34%, as measured 30 and 60 min after the distension respectively, compared with control. Pretreatment with systemic capsaicin prevented both the brain increase in Fos expression and the inhibition of gastric emptying induced by the colon distension. These results show that visceral pain arising from the proximal colon activates a complex neuronal network that includes specific brain nuclei involved in the integration of autonomic, neuroendocrine and behavioral responses to pain and an inhibitory motor reflex in other gut areas (delayed gastric emptying). Capsaicin-sensitive afferent pathways are involved in mediating brain neuronal activation and functional changes associated with noxious visceral stimulation.

Ghrelin is a newly discovered peptide hormone produced by the stomach that displays potent growth hormone-releasing activity and a stimulatory effect on food intake and digestive function while reducing energy expenditure. The isolation of ghrelin has led to new insights into how this gastric hormone links the endocrine control of nutritional homeostasis with growth hormone secretion and gastrointestinal motility through gut-brain interactions.

High-protein diet (HPD) curtails obesity and/or fat mass, but it is unknown whether it reverses neuroinflammation or alters glucose levels, CCK sensitivity, and gut microbiome in rats fed a Western diet (WD)-induced obesity (DIO). Male rats fed a WD (high fat and sugar) for 12 wk were switched to a HPD for 6 wk. Body composition, food intake, meal pattern, sensitivity to intraperitoneal CCK-8S, blood glucose, brain signaling, and cecal microbiota were assessed. When compared with a normal diet, WD increased body weight (9.3%) and fat mass (73.4%). CCK-8S (1.8 or 5.2 nmol/kg) did not alter food intake and meal pattern in DIO rats. Switching to a HPD for 6 wk reduced fat mass (15.7%) with a nonsignificantly reduced body weight gain, normalized blood glucose, and decreased feeding after CCK-8S. DIO rats on the WD or switched to a HPD showed comparable microbial diversity. However, in HPD versus WD rats, there was enrichment of 114 operational taxonomic units (OTUs) and depletion of 188 OTUs. Of those, Akkermansia muciniphila (enriched on a HPD), an unclassified Clostridiales, a member of the RF39 order, and a Phascolarctobacterium were significantly associated with fat mass. The WD increased cytokine expression in the hypothalamus and dorsal medulla that was unchanged by switching to HPD. These data indicate that HPD reduces body fat and restores glucose homeostasis and CCK sensitivity, while not modifying brain inflammation. In addition, expansion of cecal Akkermansia muciniphila correlated to fat mass loss may represent a potential peripheral mechanism of HPD beneficial effects.

The autonomic nervous system (ANS) conveys neuronal input from the brain to the stomach. We investigated mechanisms through which urocortin 1 (UCN1) injected intracerebroventricularly (ICV, 300 pmol/rat) inhibits circulating ghrelin in rats. This was achieved by assessing (1) the induction of c-fos gene expression as a marker of neuronal activation in specific hypothalamic and caudal brainstem regulating ANS; (2) the influence of vagotomy and pharmacological blockade of central and peripheral α- and β-adrenergic receptor (AR) on ICV UCN1-induced reduction of plasma ghrelin levels (determined by ELISA); and (3) the relevance of this pathway in the feeding response to a fast in rats. UCN1 increased c-fos mRNA expression in key brain sites influencing sympathetic activity namely the hypothalamic paraventricular and ventromedial nuclei, locus coeruleus, nucleus of the solitary tract, and rostral ventrolateral medulla, by 16-, 29-, 6-, 37-, and 13-fold, respectively. In contrast, the dorsal motor nucleus of the vagus had little c-fos mRNA expression and ICV UCN1 induced a similar reduction in acylated ghrelin in the sham-operated (31%) and vagotomized (41%) rats. An intraperitoneal (IP) injection of either a non-selective α- or selective α2-AR antagonist reduced, while a selective α2-AR agonist enhanced ICV UCN1-induced suppression of plasma acylated ghrelin levels. In addition, IP injection of a non-selective β- or selective β1-AR agonist blocked, and selective β1-AR antagonist augmented, the ghrelin response to ICV UCN1. The IP injections of a selective α1- or non-selective β or β2-AR antagonists, or any of the pretreatments given ICV had no effect. ICV UCN1 reduced the 2-h food intake in response to a fast by 80%, and this effect was partially prevented by a selective α2-AR antagonist. These data suggest that ICV UCN1 reduces plasma ghrelin mainly through the brain sympathetic component of the ANS and peripheral AR specifically α2-AR activation and inactivation of β1-AR. The α2-AR pathway contributes to the associated reduction in food intake.

Recently characterized selective agonists and developed antagonists for the corticotropin releasing factor (CRF) receptors are new tools to investigate stress-related functional changes. The influence of mammalian CRF and related peptides injected intracerebroventricularly (I.C.V.) on gastric and colonic motility, and the CRF receptor subtypes involved and their role in colonic response to stress were studied in conscious mice. The CRF1/CRF2 agonists rat urocortin 1(rUcn 1) and rat/human CRF (r/h CRF), the preferential CRF1 agonist ovine CRF (oCRF), and the CRF2 agonist mouse (m) Ucn 2, injected I.C.V. inhibited gastric emptying and stimulated distal colonic motor function (bead transit and defecation) while oCRF9-3OH (devoid of CRF receptor affinity) showed neither effects. mUcn 2 injected peripherally had no colonic effect. The selective CRF2 antagonist astressin2-B (I.C.V.), at a 20:1 antagonist : agonist ratio, blocked I.C.V. r/hCRF and rUcn 1 induced inhibition of gastric transit and reduced that of mUcn 2, while the CRF1 antagonist NBI-35965 had no effect. By contrast, the colonic motor stimulation induced by I.C.V. r/hCRF and rUcn 1 and 1 h restraint stress were antagonized only by NBI-35965 while stimulation induced by mUcn 2 was equally blocked by both antagonists. None of the CRF antagonists injected I.C.V. alone influenced gut transit. These data establish in mice that brain CRF1 receptors mediate the stimulation of colonic transit induced by central CRF, urocortins (1 and 2) and restraint stress, while CRF2 receptors mediate the inhibitory actions of these peptides on gastric transit. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this study was to investigate the central actions of the stable pansomatostatin peptide agonist, ODT8-SST on body weight. ODT8-SST or vehicle was acutely (1μg/rat) injected or chronically infused (5μg/rat/d, 14d) intracerebroventricularly and daily food intake, body weight and composition were monitored. In lean rats, neither acute nor chronic ODT8-SST influenced daily food intake while body weight was reduced by 2.2% after acute injection and there was a 14g reduction of body weight gain after 14d compared to vehicle (p <0.01). In diet-induced obese (DIO) rats, chronic ODT8-SST increased cumulative 2-week food intake compared to vehicle (+14%, p <0.05) and also blunted body weight change (−11g, p <0.05). ODT8-SST for 14d reduced lean mass (−22g and −25g respectively, p <0.001) and total water (−19g and −22g respectively, p <0.001) in lean and DIO rats and increased fat mass in DIO (+16g, p <0.001) but not lean rats (+1g, p >0.05) compared to vehicle. In DIO rats, ODT8-SST reduced ambulatory (−27%/24h, p <0.05) and fine movements (−38%, p <0.01) which was associated with an increased positive energy balance compared to vehicle (+50g, p <0.01). Chronic central somatostatin receptor activation in lean rats reduces body weight gain and lean mass independently of food intake which is likely related to growth hormone inhibition. In DIO rats, ODT8-SST reduces lean mass but promotes food intake and fat mass, indicating differential responsiveness to somatostatin under obese conditions. [Copyright &y& Elsevier]