Endogenous brain IL-1 mediates LPS-induced anorexia and hypothalamic cytokine expression

The present study was designed to determine the role of endogenous brain interleukin (IL)-1 in the anorexic response to lipopolysaccharide (LPS). Intraperitoneal administration of LPS (5-10 microgram/mouse) induced a dramatic, but transient, decrease in food intake, associated with an enhanced expression of proinflammatory cytokine mRNA (IL-1beta, IL-6, and tumor necrosis factor-alpha) in the hypothalamus. This dose of LPS also increased plasma levels of IL-1beta. Intracerebroventricular pretreatment with IL-1 receptor antagonist (4 microgram/mouse) attenuated LPS-induced depression of food intake and totally blocked the LPS-induced enhanced expression of proinflammatory cytokine mRNA measured in the hypothalamus 1 h after treatment. In contrast, LPS-induced increases in plasma levels of IL-1beta were not altered. These findings indicate that endogenous brain IL-1 plays a pivotal role in the development of the hypothalamic cytokine response to a systemic inflammatory stimulus.     

IL-1beta and LPS induce anorexia by distinct mechanisms differentially dependent on microsomal prostaglandin E synthase-1

Recent work demonstrated that the febrile response to peripheral immune stimulation with proinflammatory cytokine IL-1beta or bacterial wall lipopolysaccharide (LPS) is mediated by induced synthesis of prostaglandin E(2) by the terminal enzyme microsomal prostaglandin E synthase-1 (mPGES-1). The present study examined whether a similar mechanism might also mediate the anorexia induced by these inflammatory agents. Transgenic mice with a deletion of the Ptges gene, which encodes mPGES-1, and wild-type controls were injected intraperitoneally with IL-1beta, LPS, or saline. Mice were free fed, and food intake was continuously monitored with an automated system for 12 h. Body weight was recorded every 24 h for 4 days. The IL-1beta induced anorexia in wild-type but not knock-out mice, and so it was almost completely dependent on mPGES-1. In contrast, LPS induced anorexia of the same magnitude in both phenotypes, and hence it was independent of mPGES-1. However, when the mice were prestarved for 22 h, LPS induced anorexia and concomitant body weight loss in the knock-out animals that was attenuated compared with the wild-type controls. These data suggest that IL-1beta and LPS induce anorexia by distinct immune-to-brain signaling pathways and that the anorexia induced by LPS is mediated by a mechanism different from the fever induced by LPS. However, nutritional state and/or motivational factors also seem to influence the pathways for immune signaling to the brain. Furthermore, both IL-1beta and LPS caused reduced meal size but not meal frequency, suggesting that both agents exerted an anhedonic effect during these experimental conditions.     

Inhibition of TNF-alpha production contributes to the attenuation of LPS-induced hypophagia by pentoxifylline

Cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) are assumed to mediate anorexia during bacterial infections. To improve our understanding of the role that these two cytokines serve in mediating infection during anorexia, we investigated the ability of pentoxifylline (PTX), a potent inhibitor of TNF-alpha production, to block the anorectic effects of the bacterial products lipopolysaccharide (LPS) and muramyl dipeptide (MDP) in rats. Intraperitoneally injected PTX (100 mg/kg body wt) completely eliminated the anorectic effect of intraperitoneally injected LPS (100 microg/kg body wt) and attenuated the anorectic effect of a higher dose of intraperitoneally injected LPS (250 microg/kg body wt). Concurrently, PTX pretreatment suppressed low-dose LPS-induced TNF-alpha production by more than 95% and IL-1beta production 39%, as measured by ELISA. Similarly, high-dose LPS-induced TNF-alpha production was reduced by approximately 90%. PTX administration also attenuated the tolerance that is normally observed with a second injection of LPS. In addition, PTX pretreatment attenuated the hypophagic effect of intraperitoneally injected MDP (2 mg/kg body wt) but had no effect on the anorectic response to intraperitoneally injected recombinant human TNF-alpha (150 ug/kg body wt). The results suggest that suppression of TNF-alpha production is sufficient to attenuate LPS- and MDP-induced anorexia. This is consistent with the hypothesis that TNF-alpha plays a major role in the anorexia associated with bacterial infection.     

CCK(2) receptor nullification attenuates lipopolysaccharide-induced sickness behavior

Systemic infection produces a highly regulated set of responses such as fever, anorexia, adipsia, inactivity, and cachexia, collectively referred to as sickness behavior. Although the expression of sickness behavior requires immune-brain communication, the mechanisms by which peripheral cytokines signal the brain are unclear. Several mechanisms have been proposed for neuroimmune communication, including the interaction of cytokines with peripheral nerves. A critical role has been ascribed to the vagus nerve in mediating sickness behavior after intraperitoneally delivered immune activation, and converging evidence suggests that this communication may involve neurochemical intermediaries afferent and/or efferent to this nerve. Mice lacking functional CCK(2/gastrin) receptors (CCK(2)KO) and wild-type (WT) controls were administered LPS (50, 500, or 2,500 microg/kg; serotype 0111:B4; ip). Results indicate a role for CCK(2) receptor activation in the initiation and maintenance of LPS-induced sickness behavior. Compared with WT controls, CCK(2)KO mice were significantly less affected by LPS on measures of body temperature, activity, body weight, and food intake, with the magnitude of effects increasing with increasing LPS dose. Although activation of CCK(2) receptors at the level of the vagus nerve cannot be excluded, a possible role for these receptors in nonvagal routes of immune-brain communication is suggested.     

Effects of temperature and neuroactive substances on hypothalamic neurones in vitro: possible implications for the induction of fever.

This paper reviews some of our findings which have shown the usefulness of in vitro methods in the study of hypothalamic neurones. (1) Membrane current analyses of dispersed neurones of the rat preoptic and anterior hypothalamus (POA) during thermal stimulation have revealed that warm-sensitive neurones are endowed with a non-inactivating Na+ channel having a high Q10 in the hyperthermic range (35-41 degrees C). (2) A brain slice study has shown that neurones in the organum vasculosum lamina terminalis (OVLT) region have much higher sensitivity to PGE2 than POA neurones. This provides further evidence of a critical role of the OVLT in translation of blood-borne cytokine signals into brain signals for fever induction. (3) Local application of IL-1 beta and IFN alpha altered the activity of thermosensitive (TS) neurones and glucose responsive (GR) neurones in vitro in an appropriate way to produce fever and anorexia. While the responses to IL-1 beta required the local release of prostaglandins, the responses to IFN alpha were found to be mediated by opioid receptor mechanisms. (4) The responses of POA TS neurones and VMH GR neurones to IL-1 beta but not those to IFN alpha, were reversibly blocked by alpha MSH, an endogenous antipyretic peptide. Thus, immune cytokines and their related neuroactive substances may affect hypothalamic TS and GR neurones thereby producing elaborately regulated changes in homeostatic functions such as thermoregulation (fever) and feeding (anorexia), which are considered as host defence responses.     

Interleukin-1 beta-induced anorexia is reversed by ghrelin

Interleukins, in particular interleukin-1β (IL-1β), reduce food intake after peripheral and central administration, which suggests that they contribute to anorexia during various infectious, neoplastic, and autoimmune diseases. On the other hand, ghrelin stimulates food intake by acting on the central nervous system (CNS) and is considered an important regulator of food intake in both rodents and humans. In the present study, we investigated if ghrelin could reverse IL-1β-induced anorexia. Intracerebroventricular (i.c.v.) injection of 15, 30 or 45 ng/μl of IL-1β caused significant suppression of food intake in 20 h fasting animals. This effect lasted for a 24 h period. Ghrelin (0.15 nmol or 1.5 nmol/μl) produced a significant increase in cumulative food intake in normally fed animals. However, it did not alter food intake in 20 h fasting animals. Central administration of ghrelin reduced the anorexic effect of IL-1β (15 ng/μl). The effect was observed 30 min after injection and lasted for the next 24 h. This study provides evidence that ghrelin is an orexigenic peptide capable of antagonizing IL-1β-induced anorexia.     

Further characterization of high mobility group box 1 (HMGB1) as a proinflammatory cytokine: central nervous system effects

High mobility group box 1 (HMGB1), an abundant, highly conserved cellular protein, is widely known as a nuclear DNA-binding protein. HMGB1 has been recently implicated as a proinflammatory cytokine because of its role as a late mediator of endotoxin lethality and ability to stimulate release of proinflammatory cytokines from monocytes. Production of central cytokines is a critical step in the pathway by which endotoxin and peripheral proinflammatory cytokines, including interleukin-1beta (IL-1) and tumor necrosis factor-alpha (TNF), produce sickness behaviors and fever. Intracerebroventricular (ICV) administration of HMGB1 has been shown to increase TNF expression in mouse brain and induce aphagia and taste aversion. Here we show that ICV injections of HMGB1 induce fever and hypothalamic IL-1 in rats. Furthermore, we show that intrathecal administration of HMGB1 produces mechanical allodynia (lowering of the response threshold to calibrated stimuli). Finally, while endotoxin (lipopolysaccharide, LPS) administration elevates IL-1 and TNF mRNA in various brain regions, HMGB1 mRNA is unchanged. It remains possible that HMGB1 protein is released in brain in response to LPS. Nonetheless, these data suggest that HMGB1 may play a role as an endogenous pyrogen and support the concept that HMGB1 has proinflammatory characteristics within the central nervous system.     

Role of hypothalamic neuropeptide Y (NPY) in the change of feeding behavior induced by repeated treatment of amphetamine

Neuropeptide Y (NPY), an orexigenic peptide, is involved in the control of food intake. Repeated administration of amphetamine (AMPH), an anorectic agent, results in an anorectic effect on day 1 and a tolerant anorectic effect on the followings. In an attempt to know the role of hypothalamic NPY in these effects of AMPH, contents of hypothalamic NPY were determined by radioimmunoassay at first. In AMPH-treated groups, the contents of hypothalamic NPY decreased rapidly on day 1 but restored gradually to the normal level on the following days as observed in repeated AMPH. An involvement of hypothalamic NPY in the feeding change of repeated AMPH can thus be considered. Moreover, daily injection of NPY antisense oligonucleotide into brain (10 microg/10 microl/day, i.c.v.) to inhibit the gene expression of hypothalamic NPY were performed at 1 hour before daily 2 mg/kg AMPH. The reversion of food intake from the anorectic level to the normal level (tolerant anorexia) was abolished by this antisense pretreatment. It is suggested that hypothalamic NPY may play a role in the change of feeding behavior induced by repeated AMPH administration.     

Role of hypothalamic interleukin-1beta (IL-1beta) in regulation of energy homeostasis by melanocortins

In the current study we sought to determine whether hypothalamic IL-1beta is regulated by melanocortin signaling and if melanocortin-induced changes in energy balance are dependent on IL-1beta. A melanocortin agonist, MTII, increased hypothalamic IL-1beta mRNA levels by two-fold, whereas a melanocortin antagonist, SHU9119, blunted lipopolysaccharide (LPS)-mediated increase of hypothalamic IL-1beta content. Pharmacological or genetic disruption of IL-1 receptor signaling prevented MTII-mediated reductions in locomotor activity, but did not reduce MTII-induced anorexia. These data suggest a potential role for central melanocortins in mediating the decrease of ambulation characteristic of the 'sickness' response.     

Central administration of a nitric oxide precursor abolishes both the hypothalamic serotonin release and the hypophagia induced by interleukin-1beta in obese Zucker rats

Serotonin-induced anorexia has long been recognized as an important part of the CNS mechanisms controlling energy balance. More recently, interleukin-1beta and nitric oxide have been suggested to influence this control, possibly through modulation of hypothalamic serotonin. The present work aimed at investigating the interaction of these systems. We addressed whether 5-HT is affected during IL-1beta-induced anorexia in obese Zucker rats and the influence of the central NO system on this IL-1beta/5-HT interaction. Using microdialysis, we observed that an intracerebroventricular injection of 10 ng IL-1beta significantly stimulated 5-HT extracellular levels in the VMH, with a peak variation of 102+/-41% above baseline. IL-1beta also significantly reduced the 4-h feeding by 33% and the 24-h feeding by 42%. Contrarily, these effects were absent when IL-1beta was injected 2 h after the i.c.v. administration of 20 microg of the NO precursor L-arginine. The results suggest that, in obese Zucker rats, activation of the serotonergic system in the medial hypothalamus participates in IL-1beta-induced anorexia. Since L-arginine, probably through NO stimulation, abolished both the anorexia and the serotonergic activation, it can be proposed that the NO system, either directly or indirectly, counteracts IL-1beta anorexia. The hypothalamic serotonergic system is likely to mediate this NO effect.     

Attenuation of lipopolysaccharide anorexia by antagonism of caudal brain stem but not forebrain GLP-1-R

The central glucagon-like peptide-1 (GLP-1) system has been implicated in the control of feeding behavior. Here we explore GLP-1 mediation of the anorexic response to administration of systemic LPS and address the relative importance of caudal brain stem and forebrain GLP-1 receptor (GLP-1-R) for the mediation of the response. Fourth-intracerebroventricular delivery of the GLP-1-R antagonist exendin-(9-39) (10 microg) did not itself affect food intake in the 24 h after injection but significantly attenuated the otherwise robust (approximately 60%) reduction in food intake obtained after LPS (100 microg/kg) treatment. This result highlights a role for caudal brain stem GLP-1-R in the mediation of LPS anorexia but does not rule out the possibility that forebrain receptors also contribute to the response. Forebrain contribution was addressed by delivery of the GLP-1-R antagonist to the third ventricle with the caudal flow of cerebrospinal fluid blocked by occlusion of the cerebral aqueduct. Exendin-(9-39) delivery thus limited to forebrain did not attenuate the anorexic response to LPS. These data suggest that LPS anorexia is mediated, in part, by release of the native peptide acting on GLP-1-R within the caudal brain stem.     

Differential roles of tumor necrosis factor-alpha and interferon-gamma in mouse hypermetabolic and anorectic responses induced by LPS

Lipopolysaccharide (LPS)-induced effects on energy balance are characterized by alterations in energy expenditure (hypermetabolism) and food intake (anorexia). To study the role of tumour necrosis factor alpha (TNF-alpha) on some of these metabolic responses to endotoxin, we have used transgenic mice expressing soluble tumour necrosis factor receptor-1 IgG fusion protein (TNFR1-IgG) as well as TNF-alpha knockout (KO), lymphotoxin-alpha (LT-alpha) KO, and interferon-gamma receptor (IFN-gamma R) KO mice. The results from TNFR1-IgG transgenic mice suggest that the hypermetabolic and anorectic responses induced by LPS are independently regulated since, in the absence of TNF-alpha or LT-alpha, the LPS-induced hypermetabolism is almost prevented but not the anorexia. The anorectic response shows the strongest association with IFN-gamma since both IFN-gamma R KO mice and mice treated with anti-IFN-gamma antibody showed marked reduction in the LPS-induced anorexia compared to other mice. IFN-gamma R KO mice also have an attenuated thermogenic response to endotoxin. Anti-Asialo GM1 antibody treatment attenuated both the hypermetabolic and anorectic responses to LPS, to an extent comparable to that observed in IFN-gamma R KO mice. This finding suggests that natural killer cells (lymphocytic subsets) may be involved in IFN-gamma production and play an important role in the metabolic alterations induced by LPS. We also showed that the hypermetabolic response of control mice is associated with an upregulation of cytokine expression within the brain and an increase in permeability of the blood brain barrier. LPS-induced anorexia appears to involve peripheral cytokine expression.     

COX-2 inhibition attenuates anorexia during systemic inflammation without impairing cytokine production

Anorexia and weight loss are frequent complications of acute and chronic infections and result from induction of cytokines, prostaglandins, and other inflammatory mediators that are critical for pathogen elimination. Selective attenuation of the hypophagic response to infection and maintenance of the production of factors essential for infection control would be a useful addition to antimicrobial therapy in the treatment of human disease. Here, we evaluate the relative contribution of cyclooxygenase (COX)-1- and COX-2-derived prostaglandins to anorexia and weight loss precipitated by systemic immune activation by lipopolysaccharide (LPS). Using COX isoform-selective pharmacological inhibitors and gene knockout mice, we found that COX-2 inhibition during LPS-induced inflammation results in preserved food intake and maintenance of body weight, whereas COX-1 inhibition results in augmented and prolonged weight loss. Regulation of neuropeptide Y, corticotropin-releasing hormone, leptin, and interleukin-6 does not change as a function of COX-2 inhibition after LPS administration. Our data implicate COX-2 inhibition as a therapeutic target to maintain nutritional status while still allowing a normal cytokine response during infection.     

Both cyclooxygenase and lipoxygenase inhibitor partially restore the anorexia by interleukin-1 beta.

Since the peripheral prostaglandin synthetizing system may at least partly involved in the anorexia that follows central interleukin-1 beta (IL-1) administration, this study was undertaken to investigate the effect of ibuprofen (ip), selective cyclooxygenase blocker and AA 861, selective lipoxygenase inhibitor, on changes of food and water intake by a single injection of IL-1 (2 micrograms/rat, ip). We demonstrated that food and water intake were suppressed by peripheral administration of IL-1. Throughout the entire observation periods, suppressed food intake was partially restored to control levels by ibuprofen, while water intake completely restored. In addition, no significant differences about water/food intake were observed in the IL-1 + ibuprofen-treated groups, respectively. In the next experiment, IL-1 induced anorexia was also partially restored to the control level following pretreatment with AA 861. These results may suggest that other mechanism including lipoxygenase blocker besides prostaglandin production may be involved in IL-1 induced anorexia.     

A novel anti-inflammatory role for spleen-derived interleukin-10 in obesity-induced hypothalamic inflammation

Obesity can be associated with systemic low-grade inflammation that contributes to obesity-related metabolic disorders. Recent studies raise the possibility that hypothalamic inflammation contributes to the pathogenesis of diet-induced obesity (DIO), while another study reported that obesity decreases the expression of pro-inflammatory cytokines in spleen. The following study examines the hypothesis that obesity suppresses the splenic synthesis of the anti-inflammatory cytokine, interleukin (IL)-10, thereby resulting in chronic hypothalamic inflammation. The results showed that due to oxidative stress or apoptosis, the synthesis of splenic IL-10 was decreased in DIO when compared with non-obesity rats. Splenectomy (SPX) accelerated DIO-induced inflammatory responses in the hypothalamus. Interestingly, SPX suppressed the DIO-induced increases in food intake and body weight and led to a hypothalamic pro-inflammatory state that was similar to that produced by DIO, indicating that hypothalamic inflammation exerts a dual effect on energy metabolism. These SPX-induced changes were inhibited by the systemic administration of IL-10. Moreover, SPX had no effect on hypothalamic inflammatory responses in IL-10-deficient mice. These data suggest that spleen-derived IL-10 plays an important role in the prevention of hypothalamic inflammation and may be a therapeutic target for the treatment of obesity and hypothalamic inflammation.     

Cytokine neurobiology: behavior and adaptive responses

Reactions of the brain to systemic LPS or IL-1 beta treatment were shown to have different thresholds, be mediated by different neurotransmitter systems, and have different mechanisms of realisation. Changes in behaviour and neurotransmitter systems activity of the hypothalamus induced by a systemic IL-1 beta treatment were shown to be mediated by its receptors in the brain. Expression of mRNA of the tumour necrosis factor was revealed in the rabbit brain following administration of high pyrogenic doses of the LPS. The data obtained corroborate the concept of the cytokines role in maintenance of the defence responses in activation of the immune system, as well as their probable role in normal mechanisms of physiological functions control.     

Brain IL-6- and PG-dependent actions of IL-1β and lipopolysaccharide in avian fever

There is no persuasive evidence of a correlation between proinflammatory cytokines and avian fever. In this study, for the first time, we use avian cytokines to investigate a role for proinflammatory cytokines in the central component of avian fever. IL-1β and IL-6 injected intracerebroventricularly into Pekin ducks (n = 8) initiated robust fevers of equal magnitude and duration, although there was a significant difference in the latency to a febrile response. In addition, the IL-1β-induced fever could be abolished with an intracerebroventricular injection of antibodies to avian IL-6 or an oral administration of a PG synthesis inhibitor. Our findings indicate the following sequence of events within the central component of the avian febrile mechanism: IL-1β gives rise to bioactive IL-6, which stimulates an accelerated synthesis of PGs, and these PGs then adjust the sensitivity of warm-sensitive neurons in the avian brain stem to mediate fever. Yet PGE? was not upregulated in the cerebrospinal fluid of ducks made febrile with LPS. We conclude that IL-1β and IL-6 may well mediate fever by instigating an accelerated synthesis of brain-derived PG, of a class other than PGE?, or that IL-6 serves as one of the terminal mediators of the avian febrile response.     

Prophylactic effects of sulforaphane on depression-like behavior and dendritic changes in mice after inflammation

Inflammation plays a role in the pathophysiology of depression. Sulforaphane (SFN), an isothiocyanate compound derived from broccoli, is a potent activator of the NF-E2-related factor-2 (Nrf2), which plays a role in inflammation. In this study, we examined whether the prevention effects of SFN in lipopolysaccharide (LPS) induced depression-like behavior in mice. Pretreatment with SFN significantly blocked an increase in the serum tumor necrosis factor-α (TNF-α) level and an increase in microglial activation of brain regions after a single administration of LPS (0.5 mg/kg). Furthermore, SFN significantly potentiated increased serum levels of IL-10 after LPS administration. In the tail-suspension test and forced swimming test, SFN significantly attenuated an increase of the immobility time after LPS administration. In addition, SFN significantly recovered to control levels for LPS-induced alterations in the proteins such as brain-derived neurotrophic factor, postsynaptic density protein 95 and AMPA receptor 1 (GluA1) and dendritic spine density in the brain regions. Finally, dietary intake of 0.1% glucoraphanin (a glucosinolate precursor of SFN) food during the juvenile and adolescence could prevent the onset of LPS-induced depression-like behaviors and dendritic spine changes in the brain regions at adulthood. In conclusion, these findings suggest that dietary intake of SFN-rich broccoli sprout has prophylactic effects on inflammation-related depressive symptoms. Therefore, supplementation of SFN-rich broccoli sprout could be prophylactic vegetable to prevent or minimize the relapse by inflammation in the remission state of depressed patients.     

Single exposure to stressors causes long-lasting, stress-dependent reduction of food intake in rats

A single exposure to severe stressors has been shown to cause anorexia in the next 24 h, but the duration of such alterations is not known. Male Sprague-Dawley rats were subjected to different stressors, and food intake was measured for several days after stress. In experiment 1, 2 h of immobilization (Imo) and lipopolysaccharide (LPS) administration (1,000 microgram/kg) caused a marked anorexia in the 24 h after stress, which persisted on poststress day 3. In experiment 2, changes in food intake after LPS and Imo were followed until total recovery. As in experiment 1, LPS caused initially a greater degree of anorexia than Imo, but normal food intake recovered much faster (poststress day 3 vs. poststress day 9). Changing the period of exposure to Imo between 20 min and 6 h (experiment 3) only slightly modified the pattern of response to the stressor. When different doses of LPS (50, 250, and 1,000 microgram/kg) were tested in experiment 4, a dose-dependent effect on food intake was observed, the greatest doses causing the most marked and lasting effect. The present results showed stressor-specific lasting changes in food intake caused by a single exposure to some stressors, the effect of a severe psychological stressor such as Imo being more lasting than that of LPS, despite a lower initial anorexia. A severe psychological stressor and a physical stressor such as LPS appear to change food intake in different ways.