Elsevier

Molecular Brain Research

Volume 128, Issue 2, 28 September 2004, Pages 150-159
Molecular Brain Research

Research report
Increases in melanin-concentrating hormone and MCH receptor levels in the hypothalamus of dietary-obese rats

https://doi.org/10.1016/j.molbrainres.2004.06.010Get rights and content

Abstract

Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide that stimulates feeding and increases body weight in rodents. We studied the role of the system in energy homeostasis and its regulation by the satiety signals, leptin and insulin. We used real-time PCR to measure the hypothalamic expression of MCH and its receptor (MCHR1) in two contrasting models of altered nutritional status, namely, obesity induced by 8 weeks' voluntary overeating and food restriction for 10 days. Diet-fed rats were stratified according to final total fat-pad mass into a ‘high fat gain’ group (HG) and ‘low fat gain' group (LG). MCH mRNA levels were increased by 31% (p>0.05) and 49% (p<0.05) in the LG and HG, respectively, compared with controls. MCHR1 mRNA levels rose by 118% in the LG (p<0.01) and 85% in the HG (p<0.01). There were significant positive correlations (p<0.05) between plasma leptin concentration and both MCH and MCHR1 mRNA levels, and between plasma insulin and MCHR1 expression. A positive correlation was also observed between MCH and MCHR1 mRNA levels (p<0.05). Food-restricted rats showed no significant alterations in the levels of either MCH mRNA or MCHR1 mRNA. In a second experiment, we measured MCH peptide levels in five discrete hypothalamic areas of dietary-obese rats. MCH concentrations were significantly increased in the arcuate nuclei of the HG (p<0.05) and the paraventricular nuclei of both the LG (p<0.05) and HG (p<0.05), compared with their lean counterparts. These results suggest that the MCH system becomes more active in dietary obesity and could be involved in enhancing appetite for palatable food. The possibility that MCH and MCHR1 expression are positively regulated by leptin and insulin, which normally inhibit feeding, is a putative explanation for how appetite for palatable food is able to override mechanisms that prevent the development of obesity.

Introduction

Melanin-concentrating hormone (MCH) is a 19-amino acid cyclic peptide originally isolated from teleost fish, in which it regulates skin pigmentation [36]. MCH is also found in the mammalian brain where it is synthesized predominantly in neurons located in the zona incerta and lateral hypothalamic area (LHA) [11], [64]. MCH-expressing neurons have extensive fields of projection, with axons terminating throughout the central nervous system [11]. The mammalian MCH gene encodes several other putative neuropeptides including neuropeptide EI (NEI), neuropeptide GE (NGE) [45], and an alternatively spliced product termed MCH-gene-overprinted-polypeptide (MGOP) [70]. The functions of these peptides are yet to be elucidated, but it has been suggested they may regulate the translation of MCH mRNA [70] or antagonize the action of MCH [12], [58].

In mammals, MCH appears to be involved in a variety of higher functions including sensory processing [43], stress responses [35], [49], and learning [42]. Recent evidence also points to a role in energy homeostasis. The LHA has long been regarded as a ‘hunger center’, as lesions here can produce temporary hypophagia and a decrease in body weight [4]. Acute injection of MCH, either intracerebroventricularly (icv) [51], [53] or directly into the paraventricular nucleus (PVN), arcuate nucleus (ARC), or dorsomedial nucleus (DMH) [1], [54], produces a transient, dose-dependent increase in food intake. Chronic central infusion of MCH increases food intake and body weight and stimulates adiposity [19], [34]. Furthermore, mice carrying a targeted deletion of the MCH gene are hypophagic, with increased metabolic activity and weight loss [61]; conversely, transgenic mice that overexpress MCH are obese and insulin-resistant [41]. The expression of MCH is regulated by fasting and by plasma leptin concentrations. MCH mRNA is overexpressed in the hypothalamus of leptin-deficient ob/ob mice which are hyperphagic and obese [51]. Fasting further increases MCH mRNA levels in this model and also in normal rodents [29], [51], [71]. Leptin replacement in ob/ob mice and fasted rats restores MCH mRNA levels to normal. Additionally, the hyperphagia elicited by the central administration of MCH is abolished by prior injection of leptin [55]. Leptin could directly target MCH neurons in the LHA or via indirect projections from areas such as the ARC [20], [21].

The first MCH receptor (MCHR1) was identified as the orphan G protein-coupled receptor SLC-1 [5], [17], [39], [40], [57], [62], which is not activated by any other known peptide [17]. MCHR1 is widely expressed in the rat brain, with high levels in the hypothalamus, thalamus, olfactory cortex, amygdala, and hippocampus [30]. The activation of MCHR1 generates diverse intracellular signaling responses via multiple G proteins [5], [17], [28], [39], [40], [57], [62]. Selective MCHR1 antagonists suppress the food intake induced by icv administration of MCH [13], [68], as well as the consumption of palatable food [13]. The expression of MCHR1 is increased in both fasted and ob/ob mice but is unchanged in MCH knockout mice [38]. It has therefore been proposed that MCHR1 is regulated by leptin and not by MCH, unlike other G protein-coupled receptors which are inversely regulated by the availability of ligands [63]. A second receptor (MCHR2) has been identified in humans, dogs, ferrets, and monkeys. It has 32% homology with MCHR1 and is predominantly expressed in the brain [3], [31], [44], [52], [56], [73].

The aim of this study was to examine further how MCH and MCHR1 expression in the rat hypothalamus is related to energy balance and to the adiposity signals, leptin and insulin. We chose two contrasting models of altered nutritional status, namely, dietary obesity and food restriction. Both states induce marked and diametrically opposite changes in leptin and insulin and also affect various hypothalamic peptides including neuropeptide Y (NPY) [14], [77], agouti-related peptide (AGRP) [27], and pro-opiomelanocortin (POMC) [14], [37]. As a positive control, we measured POMC mRNA levels in the dietary-obese and food-restricted brain samples. In a second experiment, we measured MCH peptide levels in discrete hypothalamic areas of dietary-obese rats. This is an important model because it reflects the most common form of human obesity more than the genetically obese rodents such as the ob/ob mouse or fa/fa Zucker rat. However, in situ hybridisation studies have reported that MCH mRNA levels are not significantly different in dietary-obese rats when compared to control rats, as well as rats resistant to hypercaloric and high fat diets [22], [69]. Previous studies have examined the effect of starvation on MCH and MCHR1 mRNA levels [10], [29], [50], but the effects of food restriction have not been reported.

Section snippets

Animals

Twelve-week-old male Wistar rats (Charles River, Kent, UK) were housed in groups of two or three in a room maintained on a 12-h light–dark cycle (lights on at 0700 h) at 19–22 °C. All animals had free access to water. At the end of each study, rats were sacrificed during the early light phase (1000 h) by CO2 inhalation, blood was collected, centrifuged, and the plasma stored for subsequent assay of glucose (glucose oxidase method, Boehringer Mannheim UK, Lewes, Sussex, UK), insulin

Dietary obesity

Rats presented with the highly palatable diet responded with variable hyperphagia and ultimately developed obesity ranging in intensity from moderate to severe. To examine the role of MCH in determining this altered susceptibility to the development of dietary induced obesity, the diet-fed rats were divided into a ‘high fat gain’ group (HG), which represented the top 50% of the animals, and a ‘low fat gain’ group (LG), comprising the rest (Table 2). Total fat-pad mass was significantly

MCH and MCHR1 mRNA levels in dietary obesity

In Experiment 1, we examined the effects of chronic dietary obesity and food restriction on the expression of MCH and its receptor, MCHR1, in the whole rat hypothalamus. As MCH expression is limited to the LHA, subdividing the hypothalamus into anatomical regions would not have provided any new information. Previous studies have found that MCH expression is increased in the ob/ob mouse [51] and fa/fa rat [66] and that the peptide could contribute to sustained hyperphagia in these models. Here,

Acknowledgements

We are grateful to AstraZeneca for supporting this research, and we would especially like to thank Drs. Thomas Berglindh and Gunnar Skogman for their valuable contributions.

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