Explore the vast range of titles available.

Assess direct versus indirect action(s) of ghrelin on hypothalamic NPY neurons. Electrophysiology was used to measure ion channel activity in NPY-GFP neurons in slice preparations. Ca2+ imaging was used to monitor ghrelin activation of isolated NPY GFP-labeled neurons. Immunohistochemistry was used to localize Trpm4, SUR1 and Kir6.2 in the hypothalamus. Acylated ghrelin depolarized the membrane potential (MP) of NPY-GFP neurons in brain slices. Depolarization resulted from a decreased input resistance (IR) in ~70% of neurons (15/22) or an increased IR in the remainder (7/22), consistent with the opening or closing of ion channels, respectively. Although tetrodotoxin (TTX) blockade of presynaptic action potentials reduced ghrelin-induced changes in MP and IR, ghrelin still significantly depolarized the MP and decreased IR in TTX-treated neurons, suggesting that ghrelin directly opens cation channel(s) in NPY neurons. In isolated NPY-GFP neurons, ghrelin produced a sustained rise of [Ca2+]c, with an EC50 ~110 pM. Pharmacologic studies confirmed that the direct action of ghrelin was through occupation of the growth hormone secretagogue receptor, GHS-R, and demonstrated the importance of the adenylate cyclase/cAMP/protein kinase A (PKA) and phospholipase C/inositol triphosphate (PLC/IP3) pathways as activators of 5' AMP-activated protein kinase (AMPK). Activation of isolated neurons was not affected by CNQX or TTX, but reducing [Na+]o suppressed activation, suggesting a role for Na+-permeable cation channels. SUR1 and two channel partners, Kir6.2 and Trpm4, were identified immunologically in NPY-GFP neurons in situ. The actions of SUR1 and Trpm4 modulators were informative: like ghrelin, diazoxide, a SUR1 agonist, elevated [Ca2+]c and glibenclamide, a SUR1 antagonist, partially suppressed ghrelin action, while 9-phenanthrol and flufenamic acid, selective Trpm4 antagonists, blocked ghrelin actions on isolated neurons. Ghrelin activation was unaffected by nifedipine and ω-conotoxin, inhibitors of L- and N-type Ca2+ channels, respectively, while Ni2+, mibefradil, and TTA-P2 completely or partially inhibited ghrelin action, implicating T-type Ca2+ channels. Activation was also sensitive to a spider toxin, SNX-482, at concentrations selective for R-type Ca2+ channels. Nanomolar concentrations of GABA markedly inhibited ghrelin-activation of isolated NPY-GFP neurons, consistent with chronic suppression of ghrelin action in vivo. NPY neurons express all the molecular machinery needed to respond directly to ghrelin. Consistent with recent studies, ghrelin stimulates presynaptic inputs that activate NPY-GFP neurons in situ. Ghrelin can also directly activate a depolarizing conductance. Results with isolated NPY-GFP neurons suggest the ghrelin-activated, depolarizing current is a Na+ conductance with the pharmacologic properties of SUR1/Trpm4 non-selective cation channels. In the isolated neuron model, the opening of SUR1/Trpm4 channels activates T- and SNX482-sensitive R-type voltage dependent Ca2+ channels, which could contribute to NPY neuronal activity in situ.

BackgroundLateral and medial hypothalamus (LH and MH) play important roles in energy balance. Changed hypothalamic function has been found in subjects with obesity. However, the effect of bariatric surgery on the function of the two sub-regions has been poorly investigated.MethodsThirty-eight subjects with obesity and 34 age- and sex-matched normal-weight controls were included. Seventeen of the 38 subjects underwent laparoscopic sleeve gastrectomy. Functional magnetic resonance imaging data and metabolic parameters were collected to investigate functional connectivity networks of the two hypothalamic sub-regions as well as the influence of sleeve gastrectomy on the two networks in subjects with obesity.ResultsCompared to normal-weight controls, pre-surgical subjects had increased functional connectivity (FC) in the reward region (putamen) within the LH network, and increased FC in somatosensory cortical area (insula), as well as decreased FC in the cognitive control regions (prefrontal regions) within the MH network. After the surgery, post-surgical FC of the putamen within the LH network changed towards the patterns found in the control group. Furthermore, the changes in fasting glucose before and after the surgery were associated with the changes in FC of the putamen within the LH network.ConclusionsThe FC within the LH and MH networks were changed in subjects with obesity. Part of these altered FC was rescued after the surgery.

The DMH is known to regulate brown adipose tissue (BAT) thermogenesis via projections to sympathetic premotor neurons in the raphe pallidus, but there is evidence that the periaqueductal gray (PAG) is also an important relay in the descending pathways regulating thermogenesis. The anatomical projections from the DMH to the PAG subdivisions and their function are largely elusive, and may differ per anterior–posterior level from bregma. We here aimed to investigate the anatomical projections from the DMH to the PAG along the entire anterior–posterior axis of the PAG, and to study the role of these projections in thermogenesis in Wistar rats. Anterograde channel rhodopsin viral tracing showed that the DMH projects especially to the dorsal and lateral PAG. Retrograde rabies viral tracing confirmed this, but also indicated that the PAG receives a diffuse input from the DMH and adjacent hypothalamic subregions. We aimed to study the role of the identified DMH to PAG projections in thermogenesis in conscious rats by specifically activating them using a combination of canine adenovirus‐2 (CAV2Cre) and Cre‐dependent designer receptor exclusively activated by designer drugs (DREADD) technology. Chemogenetic activation of DMH to PAG projections increased BAT temperature and core body temperature, but we cannot exclude the possibility that at least some thermogenic effects were mediated by adjacent hypothalamic subregions due to difficulties in specifically targeting the DMH and distinct subdivisions of the PAG because of diffuse virus expression. To conclude, our study shows the complexity of the anatomical and functional connection between the hypothalamus and the PAG, and some technical challenges in studying their connection. We mapped projections from the hypothalamus to the periaquaductal grey area. Chemogenetic activation of this projection increased thermogenesis in rats.

We studied the neuronal mechanisms of the various emotional and motivational states in Wistar rats with different behavioral activities. Stimulation of the ventromedial hypothalamus had more pronounced effect on the frequency of action potentials in neurons of the dorsal hippocampus than stimulation of the lateral hypothalamus. However, stimulation of the lateral hypothalamus produced stronger effect on spatial and temporal characteristics of fi ring pattern than stimulation of the ventromedial hypothalamus. Neurons of the dorsal hippocampus in active rats were less sensitive to stimulation of the lateral and ventromedial hypothalamus than in passive animals.

The effect of cadmium (Cd) in the brain has been attributed to an increase in reactive oxygen species in cells, particularly when high amounts of the metal are given. In this study we examined the effect of a low dose of Cd (7.5 μg/day) on 24-h changes in expression of redox pathway enzyme and circadian genes in rat medial basal hypothalamus (MBH). Rats receiving CdCl₂ (5 ppm in drinking water) or tap water for 1 month were killed at six different time intervals throughout a 24 h cycle. MBH mRNA levels were measured by real-time PCR analysis. In CdCl₂ treated rats a disruption of 24-h pattern of hypothalamic gene expression of nitric oxide synthase (NOS)-1 and -2, heme oxygenase (HO)-1 and -2, Mn- superoxide dismutase (SOD), catalase, glutathione peroxidase and glutathione reductase was detectable. Mean levels of MBH mRNA for HO-2, Mn-SOD and catalase augmented after Cd intake, whereas those of NOS-2 decreased. After CdCl₂ intake rats the 24-h pattern of clock gene expression in MBH seen in controls was significantly suppressed (Bmal1) or changed in phase (Per1, Per2, Cry2) while in the case of Clock significant 24-h variations were induced. The results are compatible with the view that a low amount of Cd given in tap water brought about significant changes in circadian expression of redox enzyme and clock genes in rat MBH.

Stimulation of corticotropin-releasing factor (CRF) release from the hypothalamus by interleukin 2 (IL-2) was recently demonstrated. Cytokines induce nitric oxide synthase (NOS), an enzyme that converts L-arginine into L-citrulline and nitric oxide (NO). NO is believed to be responsible for the cytotoxic action of these agents. The constitutive form of NOS occurs in neurons in the central nervous system and NO appears to play a neurotransmitter role in cerebellar and hippocampal function. We explored the probability that IL-2 and synaptic transmitters might release CRF via NO. The effects of L-arginine, the substrate for NOS, and NG-monomethyl-L-arginine (NMMA), a competitive inhibitor of NOS, on IL-2-induced CRF release were studied using mediobasal hypothalami (MBHs) incubated in vitro in Krebs-Ringer bicarbonate buffer. L-Arginine did not alter basal and IL-2-induced CRF release after 30 min of incubation but significantly elevated both basal and IL-2-induced CRF release when MBHs were incubated 30 min longer, presumably because the endogenous substrate had been depleted after the initial 30-min incubation period. In 30-min incubations, both carbachol, an acetylcholineomimetic drug, and norepinephrine stimulated CRF release. There was an additive effect of incubation of the MBHs in the presence of carbachol (10-7M) and IL-2 (10-13M). On the other hand, coincubation of MBHs with norepinephrine (10-6M) and IL-2 (10-13M) did not produce any additive effect. Addition of NMMA, an inhibitor of NOS, at 1 or 3 * 10-4M completely suppressed IL-2-induced release of CRF as well as that caused by IL-2 plus carbachol. In contrast, the release of CRF induced by norepinephrine was not blocked by 3 * 10-4M NMMA. The data indicate that IL-2 can activate constitutive NOS leading to increased NO release, which activates CRF release. It appears that NO is also involved in the release of CRF induced by carbachol but not by norepinephrine.

We have recently shown that microinjection of the superoxide dismutase mimetic tempol into the pressor region of the rostral ventrolateral medulla attenuates the cardiovascular response to mental (air-jet) stress in rabbits. In the present study, we examined the influence of tempol on the blood pressure (BP) and heart rate (HR) responses to stress in the key region of the hypothalamic defense area, the dorsomedial hypothalamus (DMH). New Zealand White rabbits were implanted with guide cannulae for microinjection into the DMH. After 2 weeks of recovery, the cardiovascular response to air-jet stress was evaluated before and after bilateral injections of equimolar doses (20 nmol) of the superoxide scavengers tempol, tiron, or 3-carbamoyl proxyl (3-CP). Microinjection of superoxide scavengers into the DMH did not alter resting BP or HR. Air-jet stress evoked a sustained increase in BP (+16 ± 2 mm Hg) and HR (+48 ± 5 beats/min). Tempol attenuated the pressor and tachycardic responses to air-jet stress by 39% ± 10% and 37% ± 8%, respectively ( P < .05), without altering stress-induced tachypnea. Similarly, tiron selectively decreased the BP and HR responses to stress by 33% ± 8% and 53% ± 13%, respectively ( P < .05). Conversely, 3-CP, which is structurally close to tempol but has a lower superoxide scavenging activity, did not alter the cardiovascular stress response, and neither did vehicle. Microinjection of tempol or tiron just outside the DMH had little effect on stress responses. This study provides first published evidence that superoxide in the DMH is important in the regulation of acute hypertensive and tachycardic responses to mental stress.

Issue Title: Pediatric Endocrinology

Oxytocin induces mating behavior in rats of both sexes. Previous experiments revealed that progesterone-induced sex behavior in ovariectomized, extrogen-primed rats was caused by release of NO from NOergic neurons that stimulated the release of luteinizing hormone-releasing hormone (LHRH). The LHRH activated brain-stem neurons that initiated the lordosis reflex. We hypothesized that oxytocin might similarly release NO in the medial basal hypothalamic region that would stimulate release of LHRH into the hypophyseal portal vessels to release luteinizing hormone. To investigate this hypothesis, medial basal hypothalamic explants were preincubated in Krebs--Ringer bicarbonate buffer for 30 min, followed by a 30-min incubation in fresh Krebs--Ringer bicarbonate buffer containing the compounds to be tested. Oxytocin stimulated LHRH release 3- to 4-fold at the lowest concentration tested (10-10M). Values remained at a plateau as the concentration was increased to 10-7M and then declined in a concentration-dependent manner, so that there was no stimulation with a concentration of 10-5M. Oxytocin (10-7M) stimulated release of prostaglandin E2into the medium, a finding consistent with a role of NO in the response. That NO indeed mediated the action of oxytocin was supported by blockade of the action of oxytocin by the competitive inhibitor of NO synthase (NOS), NG-monomethyl-L-arginine (300 μ M). Furthermore, oxytocin (10-9to 10-7M) activated NOS as measured at the end of the experiments. Oxytocin appeared to act to stimulate norepinephrine terminals in the medial basal hypothalamus, which activated NOS by α1-adrenergic receptors, because prazocine, an α1receptor blocker, inhibited the LHRH-releasing action of oxytocin. Finally, incubation of neural lobe explants with sodium nitroprusside, a NO releasor, revealed that nitroprusside (300-600 μ M, but not 900 μ M) inhibited oxytocin release. Therefore, the NO released by oxytocin also diffuses into the oxytocin neuronal endings and inhibits oxytocin release, forming a negative feedback loop. The results indicate that oxytocin is important not only in induction of mating, but also in stimulating LHRH release with subsequent luteinizing hormone discharge that plays a crucial role in reproduction.

It has not been clearly demonstrated whether the ventromedial hypothalamus regulates intestinal cell growth. Ornithine decarboxylase is a key enzyme in polyamine synthesis, which plays an important role in intestinal mucosal growth. The aim of this study was to investigate whether bilateral ventromedial hypothalamus lesions affect mucosal cell growth. This was done by evaluating ornithine decarboxylase activity and apoptosis in rat small intestines. Bilateral ventromedial hypothalamus lesions were produced by thermocoagulation, done with rats under halothane anesthesia 7 days before the experiments. Rats with lesions were pair-fed with sham-operated rats. Total (truncal) vagotomy was performed before the development of ventromedial hypothalamus lesions. Ornithine decarboxylase activity and apoptosis were evaluated in the jejunal mucosa. Ornithine decarboxylase activity in the jejunal mucosa increased significantly 1 week after the development of the bilateral ventromedial lesions, and was attenuated by truncal vagotomy. Apoptosis in the jejunal mucosa was suppressed in rats with ventromedial hypothalamus lesions. In contrast to the effect on ornithine decarboxylase activity, the truncal vagotomies had no effect on apoptosis in rats with lesions. Apoptosis increased in the sham-operated rats after 24-h and 48-h fasting. Apoptosis in the jejunal mucosa of rats with ventromedial hypothalamus lesions did not increase after 24-h fasting. After 48-h fasting, jejunal apoptosis increased in rats with lesions, but not markedly. The ventromedial hypothalamus may regulate cell growth in the intestinal mucosa partly through the vagal nerve; however, the vagal nerve was not related to intestinal apoptosis controlled by the ventromedial hypothalamus.