Melanin-concentrating Hormone Expression Research Articles

The Influences of Chrysin on Stress-Induced Changes of Melanin-Concentrating Hormone and Orexin Gene Expression in Rats

Introduction: Chrysin is a bioactive component of herbal medicines such as Passiflora incarnate, Passiflora caerulea, and Oroxylum indicum. Although evidence has demonstrated the neuroprotective, anti-inflammatory, and pain-relieving effects of chrysin, the intra-hypothalamic molecular mechanisms underlying the anxiolytic effects of chrysin are still unclear. This study aimed to explore the effects of chrysin on hypothalamic orexin and melanin-concentrating hormone (MCH) gene expression in a rat model of stress. Methods: Twenty male Wistar rats (200±10 g) were segregated into four groups (n=5). For the induction of stress, the animals were placed in the restraint cage for 2 hours. The intact and stressed groups received saline. Thirty minutes before the induction of stress, chrysin was injected into the other two groups of the stress model at a dosage of 20 or 40 µg via the third cerebral ventricle. Hypothalamic samples were removed and frozen, and the relative gene expression of orexin and MCH was measured using the real-time polymerase chain reaction technique. Results: The induction of stress significantly increased mRNA levels of orexin and MCH compared to the control rats. The mRNA levels of MCH and orexin significantly declined in rats receiving chrysin compared to the stress group. Conclusion: The inhibition of the hypothalamic MCH and orexin neuronal circuits may be involved in the preventive effects of chrysin against stressful situations. Chrysin may be a potential target to manage anxiogenic behaviors due to the down-regulation of MCH and orexin gene expression upstream the hypothalamic corticotropin-releasing hormone.

Prolactin-Releasing Peptide Contributes to Stress-Related Mood Disorders and Inhibits Sleep/Mood Regulatory Melanin-Concentrating Hormone Neurons in Rats.

Stress disorders impair sleep and quality of life; however, their pathomechanisms are unknown. Prolactin-releasing peptide (PrRP) is a stress mediator; we therefore hypothesized that PrRP may be involved in the development of stress disorders. PrRP is produced by the medullary A1/A2 noradrenaline (NA) cells, which transmit stress signals to forebrain centers, and by non-NA cells in the hypothalamic dorsomedial nucleus. We found in male rats that both PrRP and PrRP-NA cells innervate melanin-concentrating hormone (MCH) producing neurons in the dorsolateral hypothalamus (DLH). These cells serve as a key hub for regulating sleep and affective states. Ex vivo, PrRP hyperpolarized MCH neurons and further increased the hyperpolarization caused by NA. Following sleep deprivation, intracerebroventricular PrRP injection reduced the number of REM sleep-active MCH cells. PrRP expression in the dorsomedial nucleus was upregulated by sleep deprivation, while downregulated by REM sleep rebound. Both in learned helplessness paradigm and after peripheral inflammation, impaired coping with sustained stress was associated with (1) overactivation of PrRP cells, (2) PrRP protein and receptor depletion in the DLH, and (3) dysregulation of MCH expression. Exposure to stress in the PrRP-insensitive period led to increased passive coping with stress. Normal PrRP signaling, therefore, seems to protect animals against stress-related disorders. PrRP signaling in the DLH is an important component of the PrRP's action, which may be mediated by MCH neurons. Moreover, PrRP receptors were downregulated in the DLH of human suicidal victims. As stress-related mental disorders are the leading cause of suicide, our findings may have particular translational relevance.SIGNIFICANCE STATEMENT Treatment resistance to monoaminergic antidepressants is a major problem. Neuropeptides that modulate the central monoaminergic signaling are promising targets for developing alternative therapeutic strategies. We found that stress-responsive prolactin-releasing peptide (PrRP) cells innervated melanin-concentrating hormone (MCH) neurons that are crucial in the regulation of sleep and mood. PrRP inhibited MCH cell activity and enhanced the inhibitory effect evoked by noradrenaline, a classic monoamine, on MCH neurons. We observed that impaired PrRP signaling led to failure in coping with chronic/repeated stress and was associated with altered MCH expression. We found alterations of the PrRP system also in suicidal human subjects. PrRP dysfunction may underlie stress disorders, and fine-tuning MCH activity by PrRP may be an important part of the mechanism.

Regulation and neurochemical identity of melanin-concentrating hormone neurones in the preoptic area of lactating mice.

Neurones expressing the melanin-concentrating hormone (MCH) can be found in the medial preoptic area (mPOA) and ventral aspects of the periventricular preoptic nucleus of rats by mid-to-late lactation and this expression disappears after weaning. The transitory expression of MCH in the preoptic area suggests a role for these neurones in the control of the end of lactation. However, the neurochemical identity of mPOA MCH neurones and the regulatory factors that control the transient MCH expression remain largely unknown, especially in the mouse. In the present study, we showed that mice also present the transitory expression of MCH in the mPOA at late lactation. mPOA MCH cells did not colocalise significantly with markers of GABAergic (VGAT), glutamatergic (VGLUT2 and VGLUT3) or dopaminergic (tyrosine hydroxylase) neurones. mPOA MCH cells also did not express Kiss1 or oxytocin. By contrast, approximately 70% and 90% of mPOA MCH neurones colocalised with oestrogen receptor α and prolactin-induced phosphorylated signal transducer and activator of transcription 5 (STAT5), respectively. Finally, we demonstrated that the number of MCH neurones in the mPOA is significantly higher in females during the first lactation, compared to mice on the second lactation or pregnant mice during the first lactation or brain-specific STAT5 knockout mice during the first lactation. In summary, our findings indicate that MCH neurones in the mPOA of lactating mice are sensitive to oestrogens and prolactin. Thus, mPOA MCH expression is possibly influenced by hormonal variations. Furthermore, the STAT5 signalling pathway is likely involved in the regulation of MCH expression in the mPOA of lactating mice.

Molecular codes and in vitro generation of hypocretin and melanin concentrating hormone neurons

Hypocretin/orexin (HCRT) and melanin concentrating hormone (MCH) neuropeptides are exclusively produced by the lateral hypothalamus and play important roles in sleep, metabolism, reward, and motivation. Loss of HCRT (ligands or receptors) causes the sleep disorder narcolepsy with cataplexy in humans and in animal models. How these neuropeptides are produced and involved in diverse functions remain unknown. Here, we developed methods to sort and purify HCRT and MCH neurons from the mouse late embryonic hypothalamus. RNA sequencing revealed key factors of fate determination for HCRT (Peg3, Ahr1, Six6, Nr2f2, and Prrx1) and MCH (Lmx1, Gbx2, and Peg3) neurons. Loss of Peg3 in mice significantly reduces HCRT and MCH cell numbers, while knock-down of a Peg3 ortholog in zebrafish completely abolishes their expression, resulting in a 2-fold increase in sleep amount. We also found that loss of HCRT neurons in Hcrt-ataxin-3 mice results in a specific 50% decrease in another orexigenic neuropeptide, QRFP, that might explain the metabolic syndrome in narcolepsy. The transcriptome results were used to develop protocols for the production of HCRT and MCH neurons from induced pluripotent stem cells and ascorbic acid was found necessary for HCRT and BMP7 for MCH cell differentiation. Our results provide a platform to understand the development and expression of HCRT and MCH and their multiple functions in health and disease.

P114 MCH LINKS IMMUNOMETABOLSIM TO INTESTINAL INFLAMMATION

Melanin concentrating hormone (MCH) was first identified as a hypothalamic neuropeptide regulating feeding behavior and energy balance. The MCH receptor, MCHR1, is present in brain as well as in several peripheral tissues including the gastrointestinal tract where its role is yet to be established. Increased MCH and MCHR1 expression in the mucosa of patients with inflammatory bowel disease (IBD) implies a role of MCH in disease pathogenesis. Our previous studies have shown that MCH-deficient mice develop attenuated colitis, a phenotype that was recapitulated when mice were treated with an anti-MCH antibody. The exact mechanisms by which MCH exerts proinflammatory effects remain under investigation, as well as the cell types involved and have been the focus of the present study. We found MCHR1 upregulation in CD4+ T cells isolated from the lamina propria of patients with Crohn’s disease, in contrast to those derived from peripheral blood. In vitro, MCHR1 was induced upon activation of CD4+ T-cells. Furthermore, treatment with MCH of mucosal explants from patients with IBD resulted in increased cytokine secretion in the culture supernatant (IFNg, IL-17, IL-2, IL-6, IL8, MCP1, TNFa). Most importantly, metabolomics analysis of Jurkat cells, a T-cell line expressing high basal levels of MCHR1 receptor, revealed that treatment with MCH increased glutamine, glutamate and a-ketoglutarate levels 4-, 2.5-, and 3.5-fold, respectively, as well as those of acetyl-CoA (3-fold) while glycolytic intermediates remained unaltered. These findings imply engagement of a “reductive carboxylation” cycle in the presence of MCH, the fuel of which is a-ketoglutarate instead of glucose. The role of a-ketoglutarate in driving T-effector activation and differentiation is well established. In further support of these observations, stimulation with MCH resulted in upregulation of the glutamine transporter SLC1A5 that determines intracellular glutamine levels, as well those of its metabolites glutamate and a-ketoglutarate. In relation to the immune phenotype, MCH-induced metabolic shifts correlated with increased proliferation and reduced apoptosis of Jurkat cells, as well as increased IL-2 secretion. In a more relevant context, treatment of isolated CD4+ T cells with MCH under polarized conditions resulted in 70% increase of IL-17+ cells, in contrast to 32% decrease of IL-4+ and 25% decrease of Foxp3+ cells. Collectively, these findings further support a deleterious effect of aberrant MCH signaling in IBD and potentially illustrate a new target in IBD therapeutics.

P114 MCH LINKS IMMUNOMETABOLSIM TO INTESTINAL INFLAMMATION

Melanin concentrating hormone (MCH) was first identified as a hypothalamic neuropeptide regulating feeding behavior and energy balance. The MCH receptor, MCHR1, is present in brain as well as in several peripheral tissues including the gastrointestinal tract where its role is yet to be established. Increased MCH and MCHR1 expression in the mucosa of patients with inflammatory bowel disease (IBD) implies a role of MCH in disease pathogenesis. Our previous studies have shown that MCH-deficient mice develop attenuated colitis, a phenotype that was recapitulated when mice were treated with an anti-MCH antibody. The exact mechanisms by which MCH exerts proinflammatory effects remain under investigation, as well as the cell types involved and have been the focus of the present study. We found MCHR1 upregulation in CD4+ T cells isolated from the lamina propria of patients with Crohn’s disease, in contrast to those derived from peripheral blood. In vitro, MCHR1 was induced upon activation of CD4+ T-cells. Furthermore, treatment with MCH of mucosal explants from patients with IBD resulted in increased cytokine secretion in the culture supernatant (IFNg, IL-17, IL-2, IL-6, IL8, MCP1, TNFa). Most importantly, metabolomics analysis of Jurkat cells, a T-cell line expressing high basal levels of MCHR1 receptor, revealed that treatment with MCH increased glutamine, glutamate and a-ketoglutarate levels 4-, 2.5-, and 3.5-fold, respectively, as well as those of acetyl-CoA (3-fold) while glycolytic intermediates remained unaltered. These findings imply engagement of a “reductive carboxylation” cycle in the presence of MCH, the fuel of which is a-ketoglutarate instead of glucose. The role of a-ketoglutarate in driving T-effector activation and differentiation is well established. In further support of these observations, stimulation with MCH resulted in upregulation of the glutamine transporter SLC1A5 that determines intracellular glutamine levels, as well those of its metabolites glutamate and a-ketoglutarate. In relation to the immune phenotype, MCH-induced metabolic shifts correlated with increased proliferation and reduced apoptosis of Jurkat cells, as well as increased IL-2 secretion. In a more relevant context, treatment of isolated CD4+ T cells with MCH under polarized conditions resulted in 70% increase of IL-17+ cells, in contrast to 32% decrease of IL-4+ and 25% decrease of Foxp3+ cells. Collectively, these findings further support a deleterious effect of aberrant MCH signaling in IBD and potentially illustrate a new target in IBD therapeutics.

Characterization and expression of melanin-concentrating hormone (MCH) in common carp (Cyprinus carpio) during fasting and reproductive cycle.

Melanin-concentrating hormone (MCH) was initially known as a regulator of teleost skin color and possesses multiple functions in mammals, such as the regulation of energy balance and reproduction. However, the role of MCH in fish remains unclear. In the present study, a 590bp cDNA fragment of common carp (Cyprinus carpio) MCH gene was cloned. Amino acid sequence similarities with other teleost ranged from 23 to 93%. The mature MCH peptide (DTMRCMVGRVYRPCWEV) located in the C-terminal region of MCH precursor was 100% identical to that of goldfish, zebrafish, chum salmon, and rainbow trout. Tissue expression profiles showed that MCH mRNA was ubiquitously expressed throughout the brain and peripheral tissues and highly expressed in the brain and pituitary. Within the brain, MCH mRNA was expressed preponderantly in the hypothalamus. MCH mRNA expression in the hypothalamus was increased after feeding, decreased after 3, 5, or 7days fasting, and increased upon refeeding. These results suggested that MCH might have anorexigenic actions in common carp. Meanwhile, MCH gene expression varied based on reproductive cycle, which might be related to the long-term regulation of MCH in energy balance. In conclusion, our novel finding revealed that MCH was involved in the regulation of appetite and energy balance in common carp.

Feeding state and age dependent changes in melanin-concentrating hormone expression in the hypothalamus of broiler chickens.

We aimed to quantify the gene expression changes of the potent orexigenic melanin-concentrating hormone (MCH) in chicken (Gallus gallus) hypothalamus with quantitative real-time polymerase chain reaction (qPCR), and for the first time determine peptide concentrations with a novel radioimmunoassay (RIA) under different feeding status. Three different experimental conditions, namely ad libitum feeding; fasting for 24 h; fasting for 24 h and then refeeding for 2 h, were applied to study changes of the aforementioned target and its receptor (MCHR4) gene expression under different nutritional status. The relative changes of MCH and MCHR4 were also studied from 7 to 35 days of age. Expression of PMCH and MCHR4 along the gastrointestinal tract (GIT) was also investigated. We found that expression of both targets was significant in the hypothalamus, while only weak expression was detected along the GIT. Different nutritional states did not affect the PMCH and MCHR4 mRNA levels. However, fasting for 24 h had significantly increased the MCH-like immunoreactivity by 25.65%. Fasting for 24 h and then refeeding for 2 h had further significantly increased the MCH peptide concentration by 32.51%, as compared to the ad libitum state. A decreasing trend with age was observable for both, the PMCH and MCHR4 mRNA levels, and also for the MCH-like immunoreactivity. Correlation analysis did not result in a significant correlation between MCH peptide concentration and abdominal fat mass in ad libitum fed birds. In conclusion, MCH peptide concentration altered in response to 24 h fasting, which indicated that this peptide may take part in feed intake regulation of broiler chickens.

Melanin-concentrating hormone and orexin systems in rat nucleus incertus: Dual innervation, bidirectional effects on neuron activity, and differential influences on arousal and feeding

The rat nucleus incertus (NI) contains GABA/peptide-projection neurons responsive to orexin (hypocretin)/orexin receptor-2 (OX2) signalling. Melanin-concentrating hormone (MCH) and orexin neurons often innervate and influence common target areas. Therefore, we assessed the relationship between these hypothalamic peptidergic systems and rat NI, by investigating the presence of an MCH innervation and MCH receptor-1 (MCH1) expression, and neurophysiological and behavioural effects of MCH c.f. orexin-A (OXA), within the NI. We identified lateral hypothalamus (LH), perifornical and sub-zona incerta MCH neurons that innervate NI, and characterised the rostrocaudal distribution of MCH-containing fibres in NI. Single-cell RT-PCR detected MCH1 and OX2 mRNA in NI, and multiplex, fluorescent in situ hybridisation revealed distinct co-expression patterns of MCH1 and OX2 mRNA in NI neurons expressing vesicular GABA transporter (vGAT) mRNA. Patch-clamp recordings revealed 34% of NI neurons tested were hyperpolarised by MCH (1 μM), representing a distinct population from OXA-sensitive NI neurons (35%). Intra-NI OXA infusion (600 pmol) in satiated rats during the light/inactive phase produced increased locomotor activity and food (standard chow) intake, whereas intra-NI MCH infusion (600 pmol) produced only a trend for decreased locomotor activity and no effect on food intake. Furthermore, in satiated or pre-fasted rats tested during the dark/active phase, intra-NI infusion of MCH did not alter the elevated locomotor activity or higher food intake observed. However, quantification of neuropeptide-immunostaining revealed differential diurnal fluctuations in orexin and MCH trafficking to NI. Our findings identify MCH and orexin inputs onto divergent NI populations which may differentially influence arousal and motivated behaviours.

Characterization of melanin-concentrating hormone (MCH) and its receptor in chickens: Tissue expression, functional analysis, and fasting-induced up-regulation of hypothalamic MCH expression

Melanin-concentrating hormone (MCH) is a neuropeptide expressed in the brain and exerts its actions through interaction with the two known G protein-coupled receptors, namely melanin-concentrating hormone receptor 1 and 2 (MCHR1 and MCHR2) in mammals. However, the information regarding the expression and functionality of MCH and MCHR(s) remains largely unknown in birds. In this study, using RT-PCR and RACE PCR, we amplified and cloned a MCHR1-like receptor, which is named cMCHR4 according to its evolutionary origin, and a MCHR2 from chicken brain. The cloned cMCHR4 was predicted to encode a receptor of 367 amino acids, which shares high amino acid identities with MCHR4 of ducks (90%), western painted turtles (85%), and coelacanths (77%), and a comparatively low identity to human MCHR1 (58%) and MCHR2 (38%), whereas chicken MCHR2 encodes a putative C-terminally truncated receptor and is likely a pseudogene. Using cell-based luciferase reporter assays or Western blot, we further demonstrated that chicken (and duck) MCHR4 could be potently activated by chicken MCH1–19, and its activation can elevate calcium concentration and activate MAPK/ERK and cAMP/PKA signaling pathways, indicating an important role of MCHR4 in mediating MCH actions in birds. Quantitative real-time PCR revealed that both cMCH and cMCHR4 mRNA are expressed in various brain regions including the hypothalamus, and cMCH expression in the hypothalamus of 3-week-old chicks could be induced by 36-h fasting, indicating that cMCH expression is correlated with energy balance. Taken together, characterization of chicken MCH and MCHR4 will aid to uncover the conserved roles of MCH across vertebrates.

Role of REM Sleep, Melanin Concentrating Hormone and Orexin/Hypocretin Systems in the Sleep Deprivation Pre-Ischemia

Study ObjectivesSleep reduction after stroke is linked to poor recovery in patients. Conversely, a neuroprotective effect is observed in animals subjected to acute sleep deprivation (SD) before ischemia. This neuroprotection is associated with an increase of the sleep, melanin concentrating hormone (MCH) and orexin/hypocretin (OX) systems. This study aims to 1) assess the relationship between sleep and recovery; 2) test the association between MCH and OX systems with the pathological mechanisms of stroke.MethodsSprague-Dawley rats were assigned to four experimental groups: (i) SD_IS: SD performed before ischemia; (ii) IS: ischemia; (iii) SD_Sham: SD performed before sham surgery; (iv) Sham: sham surgery. EEG and EMG were recorded. The time-course of the MCH and OX gene expression was measured at 4, 12, 24 hours and 3, 4, 7 days following ischemic surgery by qRT-PCR.ResultsA reduction of infarct volume was observed in the SD_IS group, which correlated with an increase of REM sleep observed during the acute phase of stroke. Conversely, the IS group showed a reduction of REM sleep. Furthermore, ischemia induces an increase of MCH and OX systems during the acute phase of stroke, although, both systems were still increased for a long period of time only in the SD_IS group.ConclusionsOur data indicates that REM sleep may be involved in the neuroprotective effect of SD pre-ischemia, and that both MCH and OX systems were increased during the acute phase of stroke. Future studies should assess the role of REM sleep as a prognostic marker, and test MCH and OXA agonists as new treatment options in the acute phase of stroke.

Central CCL2 signaling onto MCH neurons mediates metabolic and behavioral adaptation to inflammation.

Sickness behavior defines the endocrine, autonomic, behavioral, and metabolic responses associated with infection. While inflammatory responses were suggested to be instrumental in the loss of appetite and body weight, the molecular underpinning remains unknown. Here, we show that systemic or central lipopolysaccharide (LPS) injection results in specific hypothalamic changes characterized by a precocious increase in the chemokine ligand 2 (CCL2) followed by an increase in pro-inflammatory cytokines and a decrease in the orexigenic neuropeptide melanin-concentrating hormone (MCH). We therefore hypothesized that CCL2 could be the central relay for the loss in body weight induced by the inflammatory signal LPS. We find that central delivery of CCL2 promotes neuroinflammation and the decrease in MCH and body weight. MCH neurons express CCL2 receptor and respond to CCL2 by decreasing both electrical activity and MCH release. Pharmacological or genetic inhibition of CCL2 signaling opposes the response to LPS at both molecular and physiologic levels. We conclude that CCL2 signaling onto MCH neurons represents a core mechanism that relays peripheral inflammation to sickness behavior.

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

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. 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. 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. 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.

Physical Exercise Counteracts Stress-induced Upregulation of Melanin-concentrating Hormone in the Brain and Stress-induced Persisting Anxiety-like Behaviors.

Chronic stress induces anxiety disorders, whereas physical exercise is believed to help people with clinical anxiety. In the present study, we investigated the mechanisms underlying stress-induced anxiety and its counteraction by exercise using an established animal model of anxiety. Mice treated with restraint for 2 h daily for 14 days exhibited anxiety-like behaviors, including social and nonsocial behavioral symptoms, and these behavioral impairments lasted for more than 12 weeks after the stress treatment was removed. Despite these lasting behavioral changes, wheel-running exercise treatment for 1 h daily from post-stress days 1 - 21 counteracted anxiety-like behaviors, and these anxiolytic effects of exercise persisted for more than 2 months, suggesting that anxiolytic effects of exercise stably induced. Repeated restraint treatment up-regulated the expression of the neuropeptide, melanin-concentrating hormone (MCH), in the lateral hypothalamus, hippocampus, and basolateral amygdala, the brain regions important for emotional behaviors. In an in vitro study, treatment of HT22 hippocampal cells with glucocorticoid increased MCH expression, suggesting that MCH upregulation can be initially triggered by the stress hormone, corticosterone. In contrast, post-stress treatment with wheel-running exercise reduced the stress-induced increase in MCH expression to control levels in the lateral hypothalamus, hippocampus and basolateral amygdala. Administration of an MCH receptor antagonist (SNAP94847) to stress-treated mice was therapeutic against stress-induced anxiety-like behaviors. These results suggest that repeated stress produces long-lasting anxiety-like behaviors and upregulates MCH in the brain, while exercise counteracts stress-induced MCH expression and persisting anxiety-like behaviors.

Molecular characterization of melanin-concentrating hormone (MCH) in Schizothorax prenanti: cloning, tissue distribution and role in food intake regulation.

Melanin-concentrating hormone (MCH) is a crucial neuropeptide involved in various biological functions in both mammals and fish. In this study, the full-length MCH cDNA was obtained from Schizothorax prenanti by rapid amplification of cDNA ends polymerase chain reaction. The full-length MCH cDNA contained 589 nucleotides including an open reading frame of 375 nucleotides encoding 256 amino acids. MCH mRNA was highly expressed in the brain by real-time quantitative PCR analysis. Within the brain, expression of MCH mRNA was preponderantly detected in the hypothalamus. In addition, the MCH mRNA expression in the S. prenanti hypothalamus of fed group was significantly decreased compared with the fasted group at 1 and 3h post-feeding, respectively. Furthermore, the MCH gene expression presented significant increase in the hypothalamus of fasted group compared with the fed group during long-term fasting. After re-feeding, there was a dramatic decrease in MCH mRNA expression in the hypothalamus of S. prenanti. The results indicate that the expression of MCH is affected by feeding status. Taken together, our results suggest that MCH may be involved in food intake regulation in S. prenanti.

Prenatal exposure to ethanol stimulates hypothalamic CCR2 chemokine receptor system: Possible relation to increased density of orexigenic peptide neurons and ethanol drinking in adolescent offspring

Clinical and animal studies indicate that maternal consumption of ethanol during pregnancy increases alcohol drinking in the offspring. Possible underlying mechanisms may involve orexigenic peptides, which are stimulated by prenatal ethanol exposure and themselves promote drinking. Building on evidence that ethanol stimulates neuroimmune factors such as the chemokine CCL2 that in adult rats is shown to colocalize with the orexigenic peptide, melanin-concentrating hormone (MCH) in the lateral hypothalamus (LH), the present study sought to investigate the possibility that CCL2 or its receptor CCR2 in LH is stimulated by prenatal ethanol exposure, perhaps specifically within MCH neurons. Our paradigm of intraoral administration of ethanol to pregnant rats, at low-to-moderate doses (1 or 3g/kg/day) during peak hypothalamic neurogenesis, caused in adolescent male offspring twofold increase in drinking of and preference for ethanol and reinstatement of ethanol drinking in a two-bottle choice paradigm under an intermittent access schedule. This effect of prenatal ethanol exposure was associated with an increased expression of MCH and density of MCH+ neurons in LH of preadolescent offspring. Whereas CCL2+ cells at this age were low in density and unaffected by ethanol, CCR2+ cells were dense in LH and increased by prenatal ethanol, with a large percentage (83–87%) identified as neurons and found to colocalize MCH. Prenatal ethanol also stimulated the genesis of CCR2+ and MCH+ neurons in the embryo, which co-labeled the proliferation marker, BrdU. Ethanol also increased the genesis and density of neurons that co-expressed CCR2 and MCH in LH, with triple-labeled CCR2+/MCH+/BrdU+ neurons that were absent in control rats accounting for 35% of newly generated neurons in ethanol-exposed rats. With both the chemokine and MCH systems believed to promote ethanol consumption, this greater density of CCR2+/MCH+ neurons in the LH of preadolescent rats suggests that these systems function together in promoting alcohol drinking during adolescence.

Chronic heart failure alters orexin and melanin concentrating hormone but not corticotrophin releasing hormone-related gene expression in the brain of male Lewis rats

ObjectiveThe aim of this study was to investigate the effect of chronic heart failure (HF; 16weeks post left coronary artery ligation) on the brain's orexin (ORX) and related neuropeptide systems. MethodsIndicators of cardiac function, including the percent fractional shortening (%FS) left ventricular posterior wall shortening velocity (LVPWSV) were assessed via echocardiography at 16weeks post myocardial infarction or sham treatment in male Lewis rats (n=5/group). Changes in gene expression in HF versus control (CON) groups were quantified by real-time PCR in the hypothalamus, amygdala and dorsal pons. ResultsHF significantly reduced both the %FS and LVPWSV when compared to CON animals (P<0.02). In the hypothalamus ORX gene expression was significantly reduced in HF and correlated with changes in cardiac function when compared to CON (P<0.02). No significant changes in hypothalamic ORX receptor (type 1 or type 2) gene expression were identified. Alternatively hypothalamic melanin concentrating hormone (MCH) gene expression was significantly upregulated in HF animals and negatively correlated with LVPWSV (P<0.006). In both the amygdala and dorsal pons ORX type 2 receptor expression was significantly down-regulated in HF compared to CON. ORX receptor type 1, CRH and CRH type 1 and type 2 receptor expressions were unchanged by HF in all brain regions analyzed. ConclusionThese observations support previous work demonstrating that cardiovascular disease modulates the ORX system and identify that in the case of chronic HF the ORX system is altered in parallel with changes in MCH expression but independent of any significant changes in the central CRH system. This raises the new possibility that ORX and MCH systems may play an important role in the pathophysiology of HF.

High cortisol response to adrenocorticotrophic hormone identifies ewes with reduced melanocortin signalling and increased propensity to obesity.

We have identified female sheep that have either high (HR) or low (LR) cortisol responses to adrenocorticotrophin. On a high-energy diet, HR have greater propensity to weight gain and obesity, although the underlying mechanisms remain to be determined. Hypothalamic appetite-regulating peptides (ARP) exert reciprocal effects on food intake and energy expenditure. We aimed to quantify the expression and function of ARP in LR and HR ewes (n = 4 per group). Gene expression for neuropeptide Y (NPY), agouti-related peptide (AgRP) pro-opiomelanocortin (POMC), melanin-concentrating hormone (MCH), orexin and the melanocortin receptors (MC3R and MC4R) was measured by in situ hybridisation. Expression of NPY, AgRP and POMC was similar in HR and LR, although expression of orexin, MCH, MC3R and MC4R was higher (P < 0.05) in LR. Intracerebroventricular infusions of a low dose (50 μg/h) of NPY, α-melanocyte-stimulating hormone (αMSH), orexin and MCH were performed between 10.00 h and 16.00 h in meal-fed ewes (n = 6-7 per group). Skeletal muscle and retroperitoneal (RP) fat temperatures were recorded using dataloggers. Post-prandial thermogenesis in muscle was higher (P < 0.05) in LR. There was little effect of ARP infusion on muscle or fat temperature in either group. Infusion of these doses of NPY, MCH or orexin did not stimulate food intake in meal-fed ewes, although αMSH reduced (P < 0.01) food intake in LR only. Using 24-h ARP infusions with ad lib. feeding, NPY increased (P < 0.001) food intake in both groups but αMSH was only effective in LR (P < 0.05). In summary, we show that HR are resistant to the satiety effects of αMSH and this coincides with a reduced expression of both the MC3R and MC4R in the paraventricular nucleus of the hypothalamus. We conclude that an increased propensity to obesity in HR female sheep is associated with reduced melanocortin signalling.

In vitro assessment of interactions between appetite-regulating peptides in brain of goldfish (Carassius auratus)

Orexins, apelin, melanin-concentrating hormone (MCH), neuropeptide Y (NPY) and cocaine and amphetamine regulated transcript (CART) are important appetite-regulating factors produced by the brain of both mammals and fish. These peptide systems and their target areas are widely distributed within the central nervous system. Although morphological connections between some of these systems have been demonstrated in the brain, little is known about the functional interactions between these systems, in particular in fish. In order to better understand the interactions between appetite-related peptides, the effects of in vitro treatments of hindbrain, forebrain and hypothalamus – a major feeding regulating area – fragments with MCH, apelin and orexin on the expression of MCH, apelin, orexin, CART (forms 1 and 2) and NPY were assessed. Overall, the apelin and orexin systems stimulate each other and stimulate the NPY system while inhibiting the CART system, which is consistent with the known orexigenic actions of these two peptides. The actions of MCH remain unclear: although it appears to interact positively with orexigenic systems – as it stimulates both the orexin and apelin systems and its expression is increased by apelin – it also increases the hypothalamic expression of CART2 – but not CART1 – an anorexigenic factor, and inhibits the NPY system in all brain regions examined. This study suggests that MCH, apelin, orexin, CART and NPY do influence each other within the brain of goldfish and that these interactions might differ in nature and strength according to the peptide form and the brain region considered.

Melanin-concentrating hormone expression in the rat hypothalamus is not affected in an experiment of prenatal alcohol exposure.

Alcohol consumption during pregnancy can cause a "fetal alcoholic syndrome" (FAS) in the progeny. This syndrome is characterized by important brain defects often associated to a decreased expression of the morphogenic protein sonic hedgehog (Shh). The goal of this study was to verify if a FAS could modify the differentiation of hypothalamic neurons producing MCH. Indeed, the expression of this peptide and neurons producing it are dependent of a Shh controlled genetic cascade in the embryo. To address this question, female rats received a 15% ethanol solution to drink during pregnancy and lactation. Higher abortion rate and smaller pups at birth confirmed that descendants were affected by this experimental condition. MCH expression was analyzed by RT-qPCR and immunohistochemistry in embryos taken at E11 and E13, or in pups and young adults born from control and alcoholic mothers. MCH expression level, number of MCH neurons or ratio of MCH sub-populations were not modified by our experimental conditions. However, Shh expression was significantly lover at E11 and we also observed that hindbrain serotonergic neurons were affected as reported in the literature. These findings as well as other data from the literature suggest that protective mechanisms are involved to maintain peptide expressions and differentiation of some specific neuron populations in the ventral diencephalon in surviving embryos exposed to ethanol during pregnancy.