Rapid sensing of circulating ghrelin by hypothalamic appetite-modifying neurons

Abstract

To maintain homeostasis, hypothalamic neurons in the arcuate nucleus must dynamically sense and integrate a multitude of peripheral signals. Blood-borne molecules must therefore be able to circumvent the tightly sealed vasculature of the blood-brain barrier to rapidly access their target neurons. However, how information encoded by circulating appetite-modifying hormones is conveyed to central hypothalamic neurons remains largely unexplored. Using in vivo multiphoton microscopy together with fluorescently labeled ligands, we demonstrate that circulating ghrelin, a versatile regulator of energy expenditure and feeding behavior, rapidly binds neurons in the vicinity of fenestrated capillaries, and that the number of labeled cell bodies varies with feeding status. Thus, by virtue of its vascular connections, the hypothalamus is able to directly sense peripheral signals, modifying energy status accordingly.hormone diffusion | in vivo imaging | median eminence | metabolism C ontinuous integration of peripheral signals by neurons belonging to the arcuate nucleus of the hypothalamus (ARH) is critical for central regulation of energy balance and neuroendocrine function (1). To dynamically report alterations to homeostasis and ensure an appropriate neuronal response, blood-borne factors such as hormones must rapidly access the central nervous system (CNS). This is particularly evident in the case of food intake, which is regulated by a plethora of circulating satiety signals (2) whose levels fluctuate in an ultradian manner. Despite this, it remains unclear how key energy status-signaling hormones such as ghrelin can be rapidly sensed by target neurons to alter feeding responses (3). Elucidation of the mechanisms underlying molecule entry into the brain is important for understanding not only normal maintenance of homeostasis but also how this is perturbed during common pathologies such as obesity and diabetes (4, 5).Although molecule transport mechanisms within the ARH are poorly characterized, they likely assume one of two forms. First, chronic feedback may be accomplished by uptake of circulating molecules into the ARH via saturable receptor-mediated transport at the level of the choroid plexus and/or bloodbrain barrier (BBB) (6-9). Second, the ARH is morphologically located in close apposition to the median eminence (ME), a circumventricular organ composed of fenestrated capillaries. Because these vessels project toward the ventromedial ARH (vmARH), they could represent a direct vascular input for passive diffusion of peripheral molecules into the hypothalamus (10-13). So far, study of the functional importance of fenestrated capillaries in molecule entry into the metabolic brain has been impeded by lack of appropriate tools.To evaluate the role of fenestrated ME/ARH capillaries in rapid detection of peripheral signals by the hypothalamus, we used a recently developed in vivo imaging approach to visualize in real time the extravasation of fluorescent molecules (14). Ghrelin was chosen as a candidate hormone because i...

Palatability Can Drive Feeding Independent of AgRP Neurons

“…We reproduced our previous observation that neonatal ablation of AgRP neurons does not affect daily chow intake (see Figure S1A); however, feeding responses to peripherally or centrally injected ghrelin were abolished (See Figure S1B, S1C) (Joly-Amado et al, 2012;Luquet et al, 2005;Luquet et al, 2007). Peripheral injection of bio-active, fluorescently labeled ghrelin (Leyris et al, 2011;Schaeffer et al, 2013) resulted in robust accumulation of fluorescence in the ARC, while this signal was virtually absent in mice lacking AgRP neurons ( Figure 1A). In addition, ghrelin induced Fos in ARC neurons following intracerebroventricular (ICV) injection, which was lost in AgRP-ablated mice ( Figure 1B).…”

“…Bioactive fluorescent ghrelin was obtained as described (Schaeffer et al, 2013). Briefly, active fluorescent ghrelin (25 nmole in 100 µl) was injected i.v.…”

“…Terminally anesthetized mice at 5-10 min post-injection were perfused via the heart with 10 mL of potassium phosphate buffer 0.1 M (PBS). Brains were collected and immersed for 24 h in 4% paraformaldehyde solution (PFA) and subsequently sliced using a Leica vibratome (50-µm coronal slices) and prepared for confocal imaging as described in supplemental methods (Schaeffer et al, 2013).…”

Plasticité de la barrière hémato-hypothalamique

“…Nos résultats montrent qu'en effet l'accès de signaux métaboliques au noyau arqué est augmenté lors d'un jeûne de 24 h chez la souris [8,9]. Ceci résulte d'une modification structurale des vaisseaux du noyau arqué.…”

“…Cette barrière tanycytaire empêche les molécules d'accéder au liquide céphalo rachidien, et donc de diffuser dans le reste du cerveau, préservant l'homéos-tasie cérébrale. En effet, après le jeûne, nous observons une augmentation du taux de ghréline -molécule orexigène synthétisée par l'estomac -dans le noyau arqué ainsi que la stimulation des neurones cibles de cette molécule [9]. Ce phénomène explique le pic d'hyperphagie observé lors de la réalimentation des animaux, dont le but est de rétablir la balance énergétique.…”