Leptin sensitive neurons in the hypothalamus.

A major paradigm in the field of obesity research is the existence of an adipose tissue-brain endocrine axis for the regulation of body weight. Leptin, the peptide mediator of this axis, is secreted by adipose cells. It lowers food intake and body weight by acting in the hypothalamus, a region expressing an abundance of leptin receptors and a variety of neuropeptides that influence food intake and energy balance. Among the most promising candidates for leptin-sensitive cells in the hypothalamus are arcuate nucleus neurons that co-express the anabolic neuropeptides, neuropeptide Y (NPY) and agouti-related peptide (AGRP), and those that express proopiomelanocortin (POMC), the precursor of the catabolic peptide, alphaMSH. These cell types contain mRNA encoding leptin receptors and show changes in neuropeptide gene expression in response to changes in food intake and circulating leptin levels. Decreased leptin signaling in the arcuate nucleus is hypothesized to increase the expression of NPY and AGRP. Levels of leptin receptor mRNA and leptin binding are increased in the arcuate nucleus during fasting, principally in NPY/AGRP neurons. These findings suggest that changes in leptin receptor expression in the arcuate nucleus are inversely associated with changes in leptin signaling, and that the arcuate nucleus is an important target of leptin action in the brain.     

Somato-Dendritic Localization and Signaling by Leptin Receptors in Hypothalamic POMC and AgRP Neurons

Leptin acts via neuronal leptin receptors to control energy balance. Hypothalamic pro-opiomelanocortin (POMC) and agouti-related peptide (AgRP)/Neuropeptide Y (NPY)/GABA neurons produce anorexigenic and orexigenic neuropeptides and neurotransmitters, and express the long signaling form of the leptin receptor (LepRb). Despite progress in the understanding of LepRb signaling and function, the sub-cellular localization of LepRb in target neurons has not been determined, primarily due to lack of sensitive anti-LepRb antibodies. Here we applied light microscopy (LM), confocal-laser scanning microscopy (CLSM), and electron microscopy (EM) to investigate LepRb localization and signaling in mice expressing a HA-tagged LepRb selectively in POMC or AgRP/NPY/GABA neurons. We report that LepRb receptors exhibit a somato-dendritic expression pattern. We further show that LepRb activates STAT3 phosphorylation in neuronal fibers within several hypothalamic and hindbrain nuclei of wild-type mice and rats, and specifically in dendrites of arcuate POMC and AgRP/NPY/GABA neurons of Leprb ^+/+ mice and in Leprb ^db/db mice expressing HA-LepRb in a neuron specific manner. We did not find evidence of LepRb localization or STAT3-signaling in axon-fibers or nerve-terminals of POMC and AgRP/NPY/GABA neurons. Three-dimensional serial EM-reconstruction of dendritic segments from POMC and AgRP/NPY/GABA neurons indicates a high density of shaft synapses. In addition, we found that the leptin activates STAT3 signaling in proximity to synapses on POMC and AgRP/NPY/GABA dendritic shafts. Taken together, these data suggest that the signaling-form of the leptin receptor exhibits a somato-dendritic expression pattern in POMC and AgRP/NPY/GABA neurons. Dendritic LepRb signaling may therefore play an important role in leptin’s central effects on energy balance, possibly through modulation of synaptic activity via post-synaptic mechanisms.     

Leptin differentially regulates NPY and POMC neurons projecting to the lateral hypothalamic area.

Recent studies have reinforced the view that the lateral hypothalamic area (LHA) regulates food intake and body weight. We identified leptin-sensitive neurons in the arcuate nucleus of the hypothalamus (Arc) that innervate the LHA using retrograde tracing with leptin administration. We found that retrogradely labeled cells in the Arc contained neuropeptide Y (NPY) mRNA or proopiomelanocortin (POMC) mRNA. Following leptin administration, NPY cells in the Arc did not express Fos but expressed suppressor of cytokine signaling-3 (SOCS-3) mRNA. In contrast, leptin induced both Fos and SOCS-3 expression in POMC neurons, many of which also innervated the LHA. These findings suggest that leptin directly and differentially engages NPY and POMC neurons that project to the LHA, linking circulating leptin and neurons that regulate feeding behavior and body weight homeostasis.     

Hypothalamic NPY,AGRP, and POMC mRNA responses to leptin and refeeding in mice

Food deprivation (FD) increases hypothalamic neuropeptide Y (NPY) and agouti-related protein (AGRP) mRNA levels and decreases proopiomelanocortin (POMC) mRNA levels; refeeding restores these levels. We determined the time course of changes in hypothalamic NPY, AGRP, and POMC mRNA levels on refeeding after 24 h FD in C57BL mice by in situ hybridization. After 24 h deprivation, mice were refed with either chow or a palatable mash containing no calories or were injected with murine leptin (100 microg) without food. Mice were perfused 2 or 6 h after treatment. Food deprivation increased hypothalamic NPY mRNA (108 +/- 6%) and AGRP mRNA (78 +/- 7%) and decreased hypothalamic POMC mRNA (-15 +/- 1%). Refeeding for 6 h, but not 2 h, was sufficient to reduce (but not restore) NPY mRNA, did not affect AGRP mRNA, and restored POMC mRNA levels to ad libitum control levels. Intake of the noncaloric mash had no effect on mRNA levels, and leptin administration after deprivation (at a dose sufficient to reduce refeeding in FD mice) was not sufficient to affect mRNA levels. These results suggest that gradual postabsorptive events subsequent to refeeding are required for the restoration of peptide mRNA to baseline levels after food deprivation in mice.     

Neuropeptides,food intake and body weight regulation: a hypothalamic focus

Energy homeostasis is controlled by a complex neuroendocrine system consisting of peripheral signals like leptin and central signals, in particular, neuropeptides. Several neuropeptides with anorexigenic (POMC, CART, and CRH) as well as orexigenic (NPY, AgRP, and MCH) actions are involved in this complex (partly redundant) controlling system. Starvation as well as overfeeding lead to changes in expression levels of these neuropeptides, which act downstream of leptin, resulting in a physiological response. In this review the role of several anorexigenic and orexigenic (hypothalamic) neuropeptides on food intake and body weight regulation is summarized.     

The distribution and mechanism of action of ghrelin in the CNS demonstrates a novel hypothalamic circuit regulating energy homeostasis

The gastrointestinal peptide hormone ghrelin stimulates appetite in rodents and humans via hypothalamic actions. We discovered expression of ghrelin in a previously uncharacterized group of neurons adjacent to the third ventricle between the dorsal, ventral, paraventricular, and arcuate hypothalamic nuclei. These neurons send efferents onto key hypothalamic circuits, including those producing neuropeptide Y (NPY), Agouti-related protein (AGRP), proopiomelanocortin (POMC) products, and corticotropin-releasing hormone (CRH). Within the hypothalamus, ghrelin bound mostly on presynaptic terminals of NPY neurons. Using electrophysiological recordings, we found that ghrelin stimulated the activity of arcuate NPY neurons and mimicked the effect of NPY in the paraventricular nucleus of the hypothalamus (PVH). We propose that at these sites, release of ghrelin may stimulate the release of orexigenic peptides and neurotransmitters, thus representing a novel regulatory circuit controlling energy homeostasis.     

Recombinant leptin in the hypothalamic response to late fasting

The aim of the current study was to gain further insight into the implication of leptin in the regulation of hypothalamic gene expression during long-term food deprivation with emphasis on phase 3 of fasting (P3, late protein breakdown). Among plasma parameters, glucose, non-esterified fatty acids, and insulin levels tended to be decreased by leptin infusion, whilst corticosterone levels remained unchanged. From Northern blot analysis, NPY, AGRP, and MCH mRNA gene expressions were differentially regulated during prolonged fasting in leptin-perfused rats. In comparison with fed animals, NPY, AGRP, and MCH mRNA levels in P3 rats treated with leptin either remained stable or increased slightly. Regarding anorexigenic peptides (CART and POMC) and prepro-OX, fasting with leptin induced only slight changes in gene expression. Similar data have been obtained in leptin-treated fasted rats at various doses within the physiological range. We conclude that leptin and particularly low levels of plasma leptin can reasonably be considered as a constituent of a signal triggering the fasting-induced enhanced drive for refeeding in P3.