Melanin-concentrating hormone-expressing neurons adjust slow-wave sleep dynamics to catalyze paradoxical (REM) sleep.

Study Objectives: Experimental studies over the last 15 years established a role in sleep of the tuberal hypothalamic neurons that express melanin-concentrating hormone (MCH). Controversies still remain regarding their actual contribution to both slow-wave sleep (SWS) and paradoxical sleep (PS also known as REM sleep) or PS alone. Methods: To address this point, we compared effects of chemogenetic activation and inhibition of MCH neurons on SWS and PS amounts and EEG rhythmic activities in transgenic Pmch-cre mice. Results: In agreement with recently reported optogenetic data, the activation of MCH neurons invariably facilitates PS onset and maintenance. Our chemogenetic experiments further disclose that the ultradian rhythm of SWS is also notably related to the activity of MCH neurons. We observed that the mean duration of SWS episodes is significantly extended when MCH neurons are inhibited. Conversely, when they were excited, SWS bouts were drastically shortened and depicted substantial changes in δ rhythmic activities in electroencephalographic recording likely reflecting a deeper SWS. Conclusions: According to these original findings, we propose that when MCH neurons are physiologically recruited, SWS depth is increased and the extinction of SWS episodes is accelerated, two joint physiological processes strengthening the probability for natural SWS to PS transition and likely facilitating PS onset.

Novel Neuroprotective Effects of Melanin-Concentrating Hormone in Parkinson's Disease.

Acupuncture has shown the therapeutic effect on various neurodegenerative disorders including Parkinson's disease (PD). While investigating the neuroprotective mechanism of acupuncture, we firstly found the novel function of melanin-concentrating hormone (MCH) as a potent neuroprotective candidate. Here, we explored whether hypothalamic MCH mediates the neuroprotective action of acupuncture. In addition, we aimed at evaluating the neuroprotective effects of MCH and elucidating underlying mechanism in vitro and in vivo PD models. First, we tested whether hypothalamic MCH mediates the neuroprotective effects of acupuncture by challenging MCH-R1 antagonist (i.p.) in mice PD model. We also investigated whether MCH has a beneficial role in dopaminergic neuronal protection in vitro primary midbrain and human neuronal cultures and in vivo MPTP-induced, Pitx3-/-, and A53T mutant mice PD models. Transcriptomics followed by quantitative PCR and western blot analyses were performed to reveal the neuroprotective mechanism of MCH. We first found that hypothalamic MCH biosynthesis was directly activated by acupuncture treatment and that administration of an MCH-R1 antagonist reverses the neuroprotective effects of acupuncture. A novel finding is that MCH showed a beneficial role in dopaminergic neuron protection via downstream pathways related to neuronal survival. This is the first study to suggest the novel neuroprotective action of MCH as well as the involvement of hypothalamic MCH in the acupuncture effects in PD, which holds great promise for the application of MCH in the therapy of neurodegenerative diseases.

Dynamic Regulation of Hypothalamic DMXL2, KISS1, and RFRP Expression During Postnatal Development in Non-Human Primates.

The neurobiological mechanism of puberty onset in primates is currently only partly understood. A recent study reported an important role of Dmx-like 2 (DMXL2), a gene encoding rabconnectin-3α vesicular protein, in human subjects with mental retardation and neuroendocrine impairment of reproduction. To further characterize the potential role of DMXL2 in the regulation of reproduction, we analyzed the expression of DMXL2 in hypothalami of newborn, infantile, juvenile, pubertal, and postpubertal female and male common marmoset monkeys. Additionally, as the relative hypothalamic levels of gonadotropin-inhibitory hormone (GnIH) transcript during postnatal development are unknown in primates, we also quantified messenger RNA (mRNA) levels of RFRP, a gene encoding GnIH. Moreover, the transcript levels of kisspeptin, a well-known regulator of the hypothalamic neurohormonal axis controlling reproduction, were also checked. Transcript and protein levels of DMXL2 and Kiss1 transcript levels increase from the newborn to the infantile and from the juvenile (prepubertal) to the pubertal and the postpubertal period. We also noted a clear upsurge in RFRP transcript levels in the prepubertal period. In conclusion, the hypothalamic expressions of Kiss1 and DMXL2 mRNA increase during infantile, pubertal, and adult stages compared to newborn and juvenile stages in common marmoset monkeys. In contrast, the expression of RFRP mRNA upsurges in juvenile monkeys. Further mechanistic studies are needed to characterize the potential inhibitory role of the GnIH-GPR147 signaling in the prepubertal period and the role of DMXL2 in the molecular cascade regulating the neuroendocrine reproductive axis in primates.

Melanin-Concentrating Hormone and Its MCH-1 Receptor: Relationship Between Effects on Alcohol and Caloric Intake.

BACKGROUND: Reward and energy homeostasis are both regulated by a network of hypothalamic neuropeptide systems. The melanin-concentrating hormone (MCH) and its MCH-1 receptor (MCH1-R) modulate alcohol intake, but it remains unknown to what extent this reflects actions on energy balance or reward. Here, we evaluated the MCH1-R in regulation of caloric intake and motivation to consume alcohol in states of escalated consumption. METHODS: Rats had intermittent access (IA) to alcohol and were divided into high- and low-drinking groups. Food and alcohol consumption was assessed after administration of an MCH1-R antagonist, GW803430. Next, GW803430 was evaluated on alcohol self-administration in protracted abstinence induced by IA in high-drinking rats. Finally, the effect of GW803430 was assessed on alcohol self-administration in acute withdrawal in rats exposed to alcohol vapor. Gene expression of MCH and MCH1-R was measured in the hypothalamus and nucleus accumbens (NAc) in both acute and protracted abstinence. RESULTS: High-drinking IA rats consumed more calories from alcohol than chow and GW803430 decreased both chow and alcohol intake. In low-drinking rats, only food intake was affected. In protracted abstinence from IA, alcohol self-administration was significantly reduced by pretreatment with GW803430 and gene expression of both MCH and the MCH1-R were dysregulated in hypothalamus and NAc. In contrast, during acute withdrawal from vapor exposure, treatment with GW803430 did not affect alcohol self-administration, and no changes in MCH or MCH1-R gene expression were observed. CONCLUSIONS: Our data suggest a dual role of MCH and the MCH1-R in regulation of alcohol intake, possibly through mechanisms involving caloric intake and reward motivation. A selective suppression of alcohol self-administration during protracted abstinence by GW803430 was observed and accompanied by adaptations in gene expression of MCH and MCH1-R. Selective suppression of escalated consumption renders the MCH1-R an attractive target for treatment of alcohol use disorders.

Suckling-induced Fos activation and melanin-concentrating hormone immunoreactivity during late lactation.

AIMS: Melanin-concentrating hormone (MCH) is implicated in the control of food intake, body weight regulation and energy homeostasis. Lactation is an important physiological model to study the hypothalamic integration of peripheral sensory signals, such as suckling stimuli and those related to energy balance. MCH can be detected in the medial preoptic area (MPOA), especially around the 19th day of lactation, when this hormone is described as displaying a peak synthesis followed by a decrease after weaning. The physiological significance of this phenomenon is unclear. Therefore, we aimed to investigate hypothalamic changes associated to sensory stimulation by the litter, in special its influence over MCH synthesis. MAIN METHODS: Female Wistar rats (n=56) were euthanized everyday from lactation days 15-21, with or without suckling stimulus (WS and NS groups, respectively). MCH and Fos immunoreactivity were evaluated in the MPOA and lateral and incerto-hypothalamic areas (LHA and IHy). KEY FINDINGS: Suckling stimulus induced Fos synthesis in all regions studied. An increase on the number of suckling-induced Fos-ir neurons could be detected in the LHA after the 18th day. Conversely, the amount of MCH decreased in the MPOA from days 15-21, independent of suckling stimulation. No colocalization between MCH and Fos could be detected in any region analyzed. SIGNIFICANCE: Suckling stimulus is capable of stimulating hypothalamic regions not linked to maternal behavior, possibly to mediate energy balance aspects of lactation. Although dams are hyperphagic before weaning, this behavioral change does not appear to be mediated by MCH.

Hypothalamic ventricular ependymal thyroid hormone deiodinases are an important element of circannual timing in the Siberian hamster (Phodopus sungorus).

Exposure to short days (SD) induces profound changes in the physiology and behaviour of Siberian hamsters, including gonadal regression and up to 30% loss in body weight. In a continuous SD environment after approximately 20 weeks, Siberian hamsters spontaneously revert to a long day (LD) phenotype, a phenomenon referred to as the photorefractory response. Previously we have identified a number of genes that are regulated by short photoperiod in the neuropil and ventricular ependymal (VE) cells of the hypothalamus, although their importance and contribution to photoperiod induced physiology is unclear. In this refractory model we hypothesised that the return to LD physiology involves reversal of SD expression levels of key hypothalamic genes to their LD values and thereby implicate genes required for LD physiology. Male Siberian hamsters were kept in either LD or SD for up to 39 weeks during which time SD hamster body weight decreased before increasing, after more than 20 weeks, back to LD values. Brain tissue was collected between 14 and 39 weeks for in situ hybridization to determine hypothalamic gene expression. In VE cells lining the third ventricle, expression of nestin, vimentin, Crbp1 and Gpr50 were down-regulated at 18 weeks in SD photoperiod, but expression was not restored to the LD level in photorefractory hamsters. Dio2, Mct8 and Tsh-r expression were altered by SD photoperiod and were fully restored, or even exceeded values found in LD hamsters in the refractory state. In hypothalamic nuclei, expression of Srif and Mc3r mRNAs was altered at 18 weeks in SD, but were similar to LD expression values in photorefractory hamsters. We conclude that in refractory hamsters not all VE cell functions are required to establish LD physiology. However, thyroid hormone signalling from ependymal cells and reversal of neuronal gene expression appear to be essential for the SD refractory response.

The vertebrate diencephalic MCH system: a versatile neuronal population in an evolving brain.

Neurons synthesizing melanin-concentrating hormone (MCH) are described in the posterior hypothalamus of all vertebrates investigated so far. However, their anatomy is very different according to species: they are small and periventricular in lampreys, cartilaginous fishes or anurans, large and neuroendocrine in bony fishes, or distributed over large regions of the lateral hypothalamus in many mammals. An analysis of their comparative anatomy alongside recent data about the development of the forebrain, suggests that although very different, MCH neurons of the caudal hypothalamus are homologous. We further hypothesize that their divergent anatomy is linked to divergence in the forebrain - in particular telencephalic evolution.

Glucocorticoids are required for meal-induced changes in the expression of hypothalamic neuropeptides.

Glucocorticoid deficiency is associated with a decrease of food intake. Orexigenic peptides, neuropeptide Y (NPY) and agouti related protein (AgRP), and the anorexigenic peptide proopiomelanocortin (POMC), expressed in the arcuate nucleus of the hypothalamus (ARC), are regulated by meal-induced signals. Orexigenic neuropeptides, melanin-concentrating hormone (MCH) and orexin, expressed in the lateral hypothalamic area (LHA), also control food intake. Thus, the present study was designed to test the hypothesis that glucocorticoids are required for changes in the expression of hypothalamic neuropeptides induced by feeding. Male Wistar rats (230-280 g) were subjected to ADX or sham surgery. ADX animals received 0.9% NaCl in the drinking water, and half of them received corticosterone in the drinking water (B: 25 mg/L, ADX+B). Six days after surgery, animals were fasted for 16 h and they were decapitated before or 2 h after refeeding for brain tissue and blood collections. Adrenalectomy decreased NPY/AgRP and POMC expression in the ARC in fasted and refed animals, respectively. Refeeding decreased NPY/AgRP and increased POMC mRNA expression in the ARC of sham and ADX+B groups, with no effects in ADX animals. The expression of MCH and orexin mRNA expression in the LHA was increased in ADX and ADX+B groups in fasted condition, however there was no effect of refeeding on the expression of MCH and orexin in the LHA in the three experimental groups. Refeeding increased plasma leptin and insulin levels in sham and ADX+B animals, with no changes in leptin concentrations in ADX group, and insulin response to feeding was lower in this group. Taken together, these data demonstrated that circulating glucocorticoids are required for meal-induced changes in NPY, AgRP and POMC mRNA expression in the ARC. The lower leptin and insulin responses to feeding may contribute to the altered hypothalamic neuropeptide expression after adrenalectomy.

Galanin knockout mice show disturbances in ethanol consumption and expression of hypothalamic peptides that stimulate ethanol intake.

BACKGROUND: There is growing evidence suggesting that hypothalamic galanin (GAL), which is known to stimulate intake of a fat-rich diet, has a role in promoting the consumption of ethanol. The present study further examined this possibility in GAL knockout (GALKO) mice. METHODS: Two groups of female and male GALKO mice, compared to wild-type (WT) controls, were trained to voluntarily drink increasing concentrations of ethanol, while maintained on lab chow and water. They were examined in terms of their daily ethanol intake and preference, acute consumption of a high-fat diet, preference for flavored solutions, and expression of different peptides shown to stimulate ethanol intake. RESULTS: In the GALKO mice compared to WT, the results revealed: (i) a 35 to 45% decrease in ethanol intake and preference, which was evident only at the highest (15%) ethanol concentration, was stronger in female than in male mice, and was seen with comparisons to littermate as well as nonlittermate WT mice; (ii) a 48% decrease in acute intake of a fat-rich diet, again stronger in female than male mice; (iii) no difference in consumption of sucrose or quinine solutions in preference tests; (iv) a total loss of GAL mRNA in the hypothalamic paraventricular nucleus (PVN) of female and male mice; and (v) a gender-specific change in mRNA levels of peptides in the perifornical lateral hypothalamus (PFLH), orexin and melanin-concentrating hormone, which are known to stimulate ethanol and food intake and were markedly decreased in females while increased in males. CONCLUSIONS: These results provide strong support for a physiological role of PVN GAL in stimulating the consumption of ethanol, as well as a fat-rich diet. Ablation of the GAL gene produced a behavioral phenotype, particularly in females, which may reflect the functional relationship of galanin to ovarian steroids. It also altered the peptides in the PFLH, with their reduced expression contributing to the larger behavioral effects observed in females and their increased expression attenuating these effects in males.

Melanin-concentrating hormone producing neurons: Activities and modulations.

Regulation of energy homeostasis in animals involves adaptation of energy intake to its loss, through a perfect regulation of feeding behavior and energy storage/expenditure. Factors from the periphery modulate brain activity in order to adjust food intake as needed. Particularly, "first order" neurons from arcuate nucleus are able to detect modifications in homeostatic parameters and to transmit information to "second order" neurons, partly located in the lateral hypothalamic area. These "second order" neurons have widespread projections throughout the brain and their proper activation leads them to a coordinated response associated to an adapted behavior. Among these neurons, melanin-concentrating hormone (MCH) expressing neurons play an integrative role of the various factors arising from periphery, first order neurons and extra-hypothalamic arousal systems neurons and modulate regulation of feeding, drinking and seeking behaviors. As regulation of MCH release is correlated to regulation of MCH neuronal activity, we focused this review on the electrophysiological properties of MCH neurons from the lateral hypothalamic area. We first reviewed the knowledge on the endogenous electrical properties of MCH neurons identified according to various criteria which are described. Then, we dealt with the modulations of the electrical activity of MCH neurons by different factors such as glucose, glutamate and GABA, peptides and hormones regulating feeding and transmitters of extra-hypothalamic arousal systems. Finally, we described the current knowledge on the modulation of MCH neuronal activity by cytokines and chemokines. Because of such regulation, MCH neurons are some of the best candidate to account for infection-induced anorexia, but also obesity.

Prolactin-releasing peptide, a possible modulator of prolactin in the euryhaline silver sea bream (Sparus sarba): A molecular study.

PRL and PrRP cDNAs have been isolated from euryhaline silver sea bream (Sparus sarba). The PRL cDNA consists of 1360bp encoding 212 amino acids whereas the PrRP cDNA contains 631bp encoding preproPrRP with 122 amino acids. The mature PrRP sequence within the preprohormone is identical to the PrRPs isolated from other fish species. PRL mRNA was uniquely expressed in sea bream pituitary but PrRP mRNA was expressed in a variety of organs and tissues including the intestines, olfactory rosette and various brain regions such as hypothalamus and pituitary. Expression levels of PRL and PrRP mRNA have been examined in sea bream adapted to different salinities (0, 6, 12, 33 and 50ppt). In the pituitary, both PRL and PrRP mRNA were significantly higher in fish adapted to low salinities (0 and 6ppt) and the expression profiles of both hormones closely paralleled each other. However, expression of hypothalamic PrRP was significantly higher in fish adapted to iso-osmotic salinity (12ppt) when pituitary PRL expression was low. The present study demonstrates, for the first time, a synchronized mRNA expression pattern between PRL and PrRP in fish pituitary but a disparity of mRNA expression levels between hypothalamic PrRP and pituitary PRL during salinity adaptation. These data suggest that PrRP may possibly act as a local modulator in pituitary rather than a hypothalamic factor for regulation of pituitary PRL expression in silver sea bream.

Glucagon-like peptide 1 excites hypocretin/orexin neurons by direct and indirect mechanisms: implications for viscera-mediated arousal.

Glucagon-like peptide 1 (GLP-1) is produced by neurons in the caudal brainstem that receive sensory information from the gut and project to several hypothalamic regions involved in arousal, interoceptive stress, and energy homeostasis. GLP-1 axons and receptors have been detected in the lateral hypothalamus, where hypocretin neurons are found. The electrophysiological actions of GLP-1 in the CNS have not been studied. Here, we explored the GLP-1 effects on GFP (green fluorescent protein)-expressing hypocretin neurons in mouse hypothalamic slices. GLP-1 receptor agonists depolarized hypocretin neurons and increased their spike frequency; the antagonist exendin (9-39) blocked this depolarization. Direct GLP-1 agonist actions on membrane potential were abolished by choline substitution for extracellular Na+, and dependent on intracellular GDP, suggesting that they were mediated by sodium-dependent conductances in a G-protein-dependent manner. In voltage clamp, the GLP-1 agonist Exn4 (exendin-4) induced an inward current that reversed near -28 mV and persisted in nominally Ca2+-free extracellular solution, consistent with a nonselective cationic conductance. GLP-1 decreased afterhyperpolarization currents. GLP-1 agonists enhanced the frequency of miniature and spontaneous EPSCs with no effect on their amplitude, suggesting presynaptic modulation of glutamate axons innervating hypocretin neurons. Paraventricular hypothalamic neurons were also directly excited by GLP-1 agonists. In contrast, GLP-1 agonists had no detectable effect on neurons that synthesize melanin-concentrating hormone (MCH). Together, our results show that GLP-1 agonists modulate the activity of hypocretin, but not MCH, neurons in the lateral hypothalamus, suggesting a role for GLP-1 in the excitation of the hypothalamic arousal system possibly initiated by activation by viscera sensory input.

A role of melanin-concentrating hormone producing neurons in the central regulation of paradoxical sleep.

BACKGROUND: Peptidergic neurons containing the melanin-concentrating hormone (MCH) and the hypocretins (or orexins) are intermingled in the zona incerta, perifornical nucleus and lateral hypothalamic area. Both types of neurons have been implicated in the integrated regulation of energy homeostasis and body weight. Hypocretin neurons have also been involved in sleep-wake regulation and narcolepsy. We therefore sought to determine whether hypocretin and MCH neurons express Fos in association with enhanced paradoxical sleep (PS or REM sleep) during the rebound following PS deprivation. Next, we compared the effect of MCH and NaCl intracerebroventricular (ICV) administrations on sleep stage quantities to further determine whether MCH neurons play an active role in PS regulation. RESULTS: Here we show that the MCH but not the hypocretin neurons are strongly active during PS, evidenced through combined hypocretin, MCH, and Fos immunostainings in three groups of rats (PS Control, PS Deprived and PS Recovery rats). Further, we show that ICV administration of MCH induces a dose-dependent increase in PS (up to 200%) and slow wave sleep (up to 70%) quantities. CONCLUSION: These results indicate that MCH is a powerful hypnogenic factor. MCH neurons might play a key role in the state of PS via their widespread projections in the central nervous system.

Hypothalamic levels of NPY, MCH, and prepro-orexin mRNA during pregnancy and lactation in the rat: role of prolactin.

Pregnancy and lactation provide excellent models of physiological hyperphagia and hyperprolactinemia. To identify possible factors associated with the increased feeding in these situations, we measured hypothalamic mRNA levels of three orexigenic neuropeptides--NPY, MCH, and orexins--in nonpregnant, pregnant, and lactating rats by in situ hybridization. NPY mRNA content in the arcuate nucleus was significantly increased during pregnancy and lactation. However, MCH and prepro-orexin expression was decreased in both states. 48 or 72 h of fasting in pregnant and lactating rats further elevated NPY mRNA levels and increased the low MCH mRNA content. Surprisingly, no effect was observed in prepro-orexin mRNA levels. Finally, we investigated the possible effect of high PRL levels on these orexigenic signals using a model of hyperprolactinemia induced by pituitary graft. NPY mRNA content was unchanged, but MCH and prepro-orexin mRNA levels were significantly decreased. Our results suggest that the increased NPY expression might be partly responsible for the hyperphagia observed during pregnancy and lactation. MCH and prepro-orexin may be involved in the adaptation of other homeostatic mechanisms and their decreased levels in these physiological settings could be mediated by the elevated circulating PRL levels.

Hypothalamic gene expression in long-term fasted rats: relationship with body fat.

Many hypothalamic neuropeptides are involved in the regulation of energy homeostasis and feeding behavior. We have investigated whether and to what extent neuropeptide Y (NPY), agouti-related protein (AGRP), melanin-concentrating hormone (MCH), and prepro-orexin (prepro-OX) as well as pro-opiomelanocortin (POMC) and cocaine and amphetamine-regulated transcript (CART) mRNA levels are affected in rat hypothalamus. An experimental model of long-term fasting rat characterized by three metabolic phases from changes in lipid and protein utilization was used. Except for prepro-OX and compared to fed group, starvation induced an increase in the orexigenic gene expressions that was much more marked in phase 3 (by 2.5-, 8.1-, and 13.5-fold for MCH, AGRP, and NPY, respectively) than in phase 2 (by about 1.5-2.2-fold as an average) of fasting. AGRP and NPY mRNA levels were inversely related to body fat content. Anorexigenic gene expression was only slightly affected at both fasting stages. We conclude that the regulation of NPY and AGRP gene expression is primarily involved during late fasting and could mediate the concomitant enhanced drive for refeeding.

Involvement of the lateral hypothalamic peptide orexin in morphine dependence and withdrawal.

The lateral hypothalamus (LH) is implicated in the behavioral actions of drugs of abuse, but the cellular and molecular basis of this role is unclear. Recent identification of neuropeptides localized in LH neurons has allowed for more specific studies of LH function. The LH-specific peptide orexin (hypocretin) has been shown to be important in arousal and sleep regulation. However, orexin cells of the LH project broadly throughout the brain such that orexin may influence other behaviors as well. In this study, we show that orexin neurons, and not nearby LH neurons expressing melanin-concentrating hormone (MCH), have mu-opioid receptors and respond to chronic morphine administration and opiate antagonist-precipitated morphine withdrawal. cAMP response element-mediated transcription is induced in a subset of orexin cells, but not MCH cells, after exposure to chronic morphine or induction of withdrawal. Additionally, c-Fos and the orexin gene itself are induced in orexin cells in the LH during morphine withdrawal. Finally, we show that orexin knock-out mice develop attenuated morphine dependence, as indicated by a less severe antagonist-precipitated withdrawal syndrome. Together, these studies support a role for the orexin system in molecular adaptations to morphine, and demonstrate dramatic differences in molecular responses among different populations of LH neurons.

Selective expression of Narp, a secreted neuronal pentraxin, in orexin neurons.

Recent studies have provided compelling evidence demonstrating that orexin (also known as hypocretin) neurons play a central role in the pathophysiology of narcolepsy. However, targeted deletion of orexin does not fully mimic the functional deficits induced by selective ablation of these neurons; implying that other secreted signaling molecules expressed in these neurons mediate key aspects of their function. In this study, we demonstrate that orexin neurons display robust expression of neuronal activity-regulated pentraxin (Narp), a secreted neuronal pentraxin, implicated in regulating clustering of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors. Furthermore, we have found that hypothalamic melanin-concentrating hormone (MCH) neurons, which form a peptidergic pathway thought to oppose the effects of the orexin system, express another neuronal pentraxin, NP1. Thus, these findings suggest that these pathways utilize neuronal pentraxins, in addition to neuropeptides, as synaptic signaling molecules.

Neurochemical phenotype of hypothalamic neurons showing Fos expression 23 h after intracranial AgRP.

Agouti-related protein (AgRP) is coexpressed with neuropeptide Y (NPY) in a population of neurons in the arcuate nucleus (ARC) of the hypothalamus and stimulates food intake for up to 7 days if injected intracerebroventricularly. The prolonged food intake stimulation does not seem to depend on continued competition at the melanocortin-4 receptor (MC4R), because the relatively specific MC4R agonist MTII regains its ability to suppress food intake 24 h after AgRP injection. Intracerebroventricular AgRP also stimulates c-Fos expression 24 h after injection in several brain areas, so the neurons exhibiting delayed Fos expression might be particularly important in feeding behavior. Thus we aimed to identify the neurochemical phenotype of some of these neurons in select hypothalamic areas, using double-label immunohistochemistry. AgRP-injected rats ingested significantly more chow (10.2 +/- 0.6 g) vs. saline controls (3.4 +/- 0.7 g) in the first 9 h (light phase) after injection. In the lateral hypothalamus (particularly the perifornical area) 23 h after injection, AgRP induced significantly more Fos vs. saline in orexin-A (OXA) neurons (25.6 +/- 4.9 vs. 4.8 +/- 3.1%), but not in melanin-concentrating hormone (MCH) or cocaine- and amphetamine-regulated transcript (CART) neurons. In the ARC, AgRP induced significantly more Fos in CART (40.6 +/- 5.9 vs. 13.4 +/- 1.8%) but not NPY neurons. In the paraventricular nucleus, there was no significant difference in Fos expression induced by AgRP vs. saline in oxytocin and CART neurons. We conclude that the long-lasting hyperphagia induced by AgRP is correlated with and possibly partially mediated by hyperactive OXA neurons in the lateral hypothalamus and CART neurons in the ARC, but not by NPY and MCH neurons. The substantial increase in light-phase food intake by AgRP supports a role for the arousing effects of OXA. Activation of CART neurons in the ARC (which likely coexpress proopiomelanocortin) could indicate attempts to activate counterregulatory decreases in food intake.

Effect of a fatty acid synthase inhibitor on food intake and expression of hypothalamic neuropeptides.

The fatty acid synthase inhibitor, C75, acts centrally to reduce food intake and body weight in mice. Here we report the effects of C75 on the expression of key orexigenic [neuropeptide Y (NPY), agouti-related protein (AgRP), and melanin-concentrating hormone] and anorexigenic [pro-opiomelanocortin (POMC) and cocaine-amphetamine-related transcript (CART)] neuropeptide messages in the hypothalami of lean and obese (ob/ob) mice. In lean mice, C75 rapidly and almost completely blocked food intake and prevented fasting-induced up-regulation of hypothalamic AgRP and NPY mRNAs, as well as down-regulation of CART and POMC mRNAs. Thus, in lean mice C75 seems to interrupt the fasting-induced signals that activate expression of NPY and AgRP and suppression of POMC and CART. In obese mice, C75 rapidly suppressed food intake, reduced body weight, and normalized obesity-associated hyperglycemia and hyperinsulinemia. Like its effect in lean mice, C75 prevented the fasting-induced increase of hypothalamic NPY and AgRP mRNAs in obese mice, but had no effect on the expression of POMC and CART mRNAs. The suppressive effect of C75 on food intake in lean mice seems to be mediated both by NPY/AgRP and POMC/CART neurons, whereas in obese mice the effect seems to be mediated primarily by NPY/AgRP neurons. In both lean and obese mice, C75 markedly increased expression of melanin-concentrating hormone and its receptor in the hypothalamus.