Elsevier

Brain Research

Volume 1119, Issue 1, 13 November 2006, Pages 101-114
Brain Research

Research Report
MCH-containing neurons in the hypothalamus of the cat: Searching for a role in the control of sleep and wakefulness

https://doi.org/10.1016/j.brainres.2006.08.100Get rights and content

Abstract

Neurons that utilize melanin-concentrating hormone (MCH) and others that employ hypocretin as neurotransmitter are located in the hypothalamus and project diffusely throughout the CNS, including areas that participate in the generation and maintenance of the states of sleep and wakefulness. In the present report, immunohistochemical methods were employed to examine the distribution of MCHergic and hypocretinergic neurons. In order to test the hypothesis that the MCHergic system is capable of influencing specific behavioral states, we studied Fos immunoreactivity in MCH-containing neurons during (1) quiet wakefulness, (2) active wakefulness with motor activity, (3) active wakefulness without motor activity, (4) quiet sleep and (5) active sleep induced by carbachol (AS-carbachol). We determined that MCHergic neuronal somata in the cat are intermingled with hypocretinergic neurons in the dorsal and lateral hypothalamus, principally in the tuberal and tuberomammillary regions; however, hypocretinergic neurons extended more in the anterior–posterior axis than MCHergic neurons. Axosomatic and axodendritic contacts were common between these neurons. In contrast to hypocretinergic neurons, which are known to be active during motor activity and AS-carbachol, Fos immunoreactivity was not observed in MCH-containing neurons in conjunction with any of the preceding behavioral conditions. Non-MCHergic, non-hypocretinergic neurons that expressed c-fos during active wakefulness with motor activity were intermingled with MCH and hypocretin-containing neurons, suggesting that these neurons are related to some aspect of motor function. Further studies are required to elucidate the functional sequela of the interactions between MCHergic and hypocretinergic neurons and the phenotype of the other neurons that were active during motor activity.

Introduction

The melanin-concentrating hormone (MCH) was initially characterized as a circulating factor mediating color change in teleost fish (Kawauchi et al., 1983). Thereafter, MCH was identified in the rat and was found to be fully conserved in all mammals analyzed so far, including humans (Forray, 2003, Shi, 2004). MCH is generated via a proteolytic cleavage of a precursor that generates two additional peptides, neuropeptide E-I and neuropeptide G-E. The biological function of MCH is mediated by two G-protein-coupled receptors known as MCH1R and MCHR2, although MCHR2 is only expressed in carnivores and primates (Forray, 2003, Shi, 2004). The conservation of this peptide across species suggests that MCH is involved in critical physiological processes. In this regard, there is evidence that MCH plays a key role in the central regulation of feeding and energy homeostasis. In genetically modified obese mice, there is an up-regulation of MCH mRNA, and MCH produces a dose-dependent increase in food intake when administered intraventricularly in rats (Qu et al., 1996, Rossi et al., 1997, Rovere et al., 1996). This stimulatory effect on food intake has been confirmed in transgenic mice in which the overexpression of MCH causes obesity, whereas mice lacking MCH are hypophagic and lean (Ludwig et al., 2001, Shimada et al., 1998). In addition, there are data indicating that the pharmacological blockade of MCH receptors may be used in a therapeutic manner to treat obesity, depression and anxiety, suggesting that the MCHergic system participates in other functions (Borowsky et al., 2002).

Neurons that utilize MCH as a neurotransmitter have been described in the lateral hypothalamus and zona incerta of the rat (Bittencourt et al., 1992, Skofitsch et al., 1985). These neurons project throughout the central nervous system with dense projections to areas associated with the regulation of sleep and wakefulness (Bittencourt et al., 1992, Jones, 2005, Skofitsch et al., 1985); high densities of MCH receptors are also present in these areas (Kilduff and De Lecea, 2001). Based upon experiments utilizing Fos immunoreactivity as a marker of neuronal activity as well as MCH intraventricular microinjections, Verret et al. (2003) suggested that MCH is a powerful hypnogenic factor that plays a crucial role in the control of active (REM) sleep (AS) in the rat. In hamsters and rats, MCHergic neurons are intermingled with hypocretinergic neurons and make numerous synaptic contacts with these cells (Bayer et al., 2002, Guan et al., 2002, Khorooshi and Klingenspor, 2005). The hypocretinergic system has an anatomical organization similar to the MCHergic system (Peyron et al., 1998), and it has been proposed to be involved in the regulation of behavioral states; deficits in this system have been associated with narcolepsy/cataplexy (Siegel, 2004, Sutcliffe and de Lecea, 2002, Taheri et al., 2002). In the present report, as a foundation for furthering understanding of the function of the MCHergic system and its interactions with the hypocretinergic system, we determined the location of the neuronal somata of MCH-containing neurons and their anatomical relationship with the hypocretinergic system in the cat. In addition, we studied Fos immunoreactivity in MCH-containing neurons during quiet and active waking as well as during quiet (NREM) sleep (QS) and AS, in order to reveal the activity of these cells vis-a-vis behavioral states.

Section snippets

Location of MCH-containing somata

Neuronal somata and processes were observed after immunostaining against MCH and/or hypocretin. MCHergic neurons were located in the hypothalamus (Fig. 1). These neurons were intermingled with and were in close proximity to hypocretinergic neurons; however, these neuropeptides were not co-localized in the same cells (Fig. 2). The absence of co-localization was observed throughout the hypothalamus.

MCHergic neurons were concentrated in the perifornical region of the lateral hypothalamic area,

Discussion

Since the pioneering work of von Economo (1930), the hypothalamus has been the object of many studies dealing with sleep physiology and pathology. Renewed interest in the hypothalamus has emerged due to the discovery of the hypocretinergic system which has been suggested to be involved in the control of motor activity, sleep and wakefulness (reviewed by Siegel, 2004, Sutcliffe and de Lecea, 2002, Taheri et al., 2002). MCHergic neurons are intermingled with the hypocretinergic neurons in the

Experimental procedures

Sixteen adult cats, which were used in the present study, were obtained from and determined to be in good health by the UCLA Division of Laboratory Animal Medicine. All experimental procedures were conducted in accord with the “Guide to the care and use of laboratory animals” (7th edition, National Academy Press, Washington, DC, 1996) and were approved by the Animal Research Committee of the UCLA Office for the Protection of the Research Subjects; adequate measures were taken to minimize pain

Acknowledgments

We thank Dr. J. K. Engelhardt for his critical comments regarding the manuscript. This study was supported by USPHS grants MH43362, MH 69372, NS09999, NS23426, HL602969 and AG04307.

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