Ontogeny of purinergic receptor-regulated Ca2+ signaling in mouse cortical collecting duct epithelium

Changes in ATP-induced increase in {[}Ca2+], during collecting duct ontogeny were studied in primary monolayer cultures of mouse ureteric bud (UB) and cortical collecting duct (CCD) cells by Fura-PE3 fluorescence ratio imaging. In UB (embryonic day E14 and postnatal day P1) the ATIP-stimulated increase (EC50 approximate to 1 muM) in fluorescence ratio (DeltaR(ATP)) was independent of extracellular Ca2+ and insensitive to the P2 purinoceptor-antagonist suramin (1 mM). From day P7 onward when CCD morphogenesis had been completed DeltaR(ATP) increased and became dependent on extracellular Ca2+. This ATP-stimulated Ca2+ entry into CCD cells was non-capacitative and suramin (11 mM)insensitive, but sensitive to nifedipine (30 muM) and enhanced by Bay K8644 (15 muM), a blocker and an agonist of L-type Ca2+ channels, respectively. Quantitative RT-PCR demonstrated similar mRNA expression of L-type Ca2+ channel alpha1-subunit, P2Y(1), P2Y(2), and P2X(4b) purinoceptors in UB and CCD monolayers while the abundance of P2X(4) mRNA increased with CCD morphogenesis. In conclusion, both embryonic and postnatal cells express probably P2Y(2)-stimulated Ca2+ release from intracellular stores. With development, the CCD epithelium acquires ATP-stimulated Ca2+ entry via L-type Ca2+ channels. This pathway might by mediated by the increasing expression of P2X(4)-receptors resulting in an increasing ATP-dependent membrane depolarization and activation of L-type Ca2+ channels. Copyright (C) 2002 S. Karger AG, Basel

Unacylated Ghrelin Promotes Skeletal Muscle Regeneration Following Hindlimb Ischemia via SOD-2-Mediated miR-221/222 Expression

BACKGROUND: Surgical treatment of peripheral artery disease, even if successful, does not prevent reoccurrence. Under these conditions, increased oxidative stress is a crucial determinant of tissue damage. Given its reported antioxidant effects, we investigated the potential of unacylated‐ghrelin (UnAG) to reduce ischemia‐induced tissue damage in a mouse model of peripheral artery disease. METHODS AND RESULTS: We show that UnAG but not acylated ghrelin (AG) induces skeletal muscle regeneration in response to ischemia via canonical p38/mitogen‐actived protein kinase signaling UnAG protected against reactive oxygen species–induced cell injuries by inducing the expression of superoxide dismutase‐2 (SOD‐2) in satellite cells. This led to a reduced number of infiltrating CD68(+) cells and was followed by induction of the myogenic process and a reduction in functional impairment. Moreover, we found that miR‐221/222, previously linked to muscle regeneration processes, was up‐regulated and negatively correlated with p57(Kip2) expression in UnAG‐treated mice. UnAG, unlike AG, promoted cell‐cycle entry in satellite cells of mice lacking the genes for ghrelin and its receptor (GHSR1a). UnAG‐induced p38/mitogen‐actived protein kinase phosphorylation, leading to activation of the myogenic process, was prevented in SOD‐2–depleted SCs. By siRNA technology, we also demonstrated that SOD‐2 is the antioxidant enzyme involved in the control of miR‐221/222–driven posttranscriptional p57(Kip2) regulation. Loss‐of‐function experiments targeting miR‐221/222 and local pre–miR‐221/222 injection in vivo confirmed a role for miR‐221/222 in driving skeletal muscle regeneration after ischemia. CONCLUSIONS: These results indicate that UnAG‐induced skeletal muscle regeneration after ischemia depends on SOD‐2–induced miR‐221/222 expression and highlight its clinical potential for the treatment of reactive oxygen species–mediated skeletal muscle damage

Preprandial ghrelin is not affected by macronutrient intake, energy intake or energy expenditure

BACKGROUND: Ghrelin, a peptide secreted by endocrine cells in the gastrointestinal tract, is a hormone purported to have a significant effect on food intake and energy balance in humans. The influence of factors related to energy balance on ghrelin, such as daily energy expenditure, energy intake, and macronutrient intake, have not been reported. Secondly, the effect of ghrelin on food intake has not been quantified under free-living conditions over a prolonged period of time. To investigate these effects, 12 men were provided with an ad libitum cafeteria-style diet for 16 weeks. The macronutrient composition of the diets were covertly modified with drinks containing 2.1 MJ of predominantly carbohydrate (Hi-CHO), protein (Hi-PRO), or fat (Hi-FAT). Total energy expenditure was measured for seven days on two separate occasions (doubly labeled water and physical activity logs). RESULTS: Preprandial ghrelin concentrations were not affected by macronutrient intake, energy expenditure or energy intake (all P > 0.05). In turn, daily energy intake was significantly influenced by energy expenditure, but not ghrelin. CONCLUSION: Preprandial ghrelin does not appear to be influenced by macronutrient composition, energy intake, or energy expenditure. Similarly, ghrelin does not appear to affect acute or chronic energy intake under free-living conditions

Ghrelin Indirectly Activates Hypophysiotropic CRF Neurons in Rodents

Ghrelin is a stomach-derived hormone that regulates food intake and neuroendocrine function by acting on its receptor, GHSR (Growth Hormone Secretagogue Receptor). Recent evidence indicates that a key function of ghrelin is to signal stress to the brain. It has been suggested that one of the potential stress-related ghrelin targets is the CRF (Corticotropin-Releasing Factor)-producing neurons of the hypothalamic paraventricular nucleus, which secrete the CRF neuropeptide into the median eminence and activate the hypothalamic-pituitary-adrenal axis. However, the neural circuits that mediate the ghrelin-induced activation of this neuroendocrine axis are mostly uncharacterized. In the current study, we characterized in vivo the mechanism by which ghrelin activates the hypophysiotropic CRF neurons in mice. We found that peripheral or intra-cerebro-ventricular administration of ghrelin strongly activates c-fos – a marker of cellular activation – in CRF-producing neurons. Also, ghrelin activates CRF gene expression in the paraventricular nucleus of the hypothalamus and the hypothalamic-pituitary-adrenal axis at peripheral level. Ghrelin administration directly into the paraventricular nucleus of the hypothalamus also induces c-fos within the CRF-producing neurons and the hypothalamic-pituitary-adrenal axis, without any significant effect on the food intake. Interestingly, dual-label immunohistochemical analysis and ghrelin binding studies failed to show GHSR expression in CRF neurons. Thus, we conclude that ghrelin activates hypophysiotropic CRF neurons, albeit indirectly

High-Fat Diet with Acyl-Ghrelin Treatment Leads to Weight Gain with Low Inflammation, High Oxidative Capacity and Normal Triglycerides in Rat Muscle

Obesity is associated with muscle lipid accumulation. Experimental models suggest that inflammatory cytokines, low mitochondrial oxidative capacity and paradoxically high insulin signaling activation favor this alteration. The gastric orexigenic hormone acylated ghrelin (A-Ghr) has antiinflammatory effects in vitro and it lowers muscle triglycerides while modulating mitochondrial oxidative capacity in lean rodents. We tested the hypothesis that A-Ghr treatment in high-fat feeding results in a model of weight gain characterized by low muscle inflammation and triglycerides with high muscle mitochondrial oxidative capacity. A-Ghr at a non-orexigenic dose (HFG: twice-daily 200-µg s.c.) or saline (HF) were administered for 4 days to rats fed a high-fat diet for one month. Compared to lean control (C) HF had higher body weight and plasma free fatty acids (FFA), and HFG partially prevented FFA elevation (P<0.05). HFG also had the lowest muscle inflammation (nuclear NFkB, tissue TNF-alpha) with mitochondrial enzyme activities higher than C (P<0.05 vs C, P = NS vs HF). Under these conditions HFG prevented the HF-associated muscle triglyceride accumulation (P<0.05). The above effects were independent of changes in redox state (total-oxidized glutathione, glutathione peroxidase activity) and were not associated with changes in phosphorylation of AKT and selected AKT targets. Ghrelin administration following high-fat feeding results in a novel model of weight gain with low inflammation, high mitochondrial enzyme activities and normalized triglycerides in skeletal muscle. These effects are independent of changes in tissue redox state and insulin signaling, and they suggest a potential positive metabolic impact of ghrelin in fat-induced obesity

Fasting and postprandial plasma ghrelin levels are decreased in patients with liver failure previous to liver transplantation

[Abstract] Anorexia is a problem of paramount importance in patients with advanced liver failure. Ghrelin has important actions on feeding and weight homeostasis. Concentrations of ghrelin are controversial in liver cirrhosis. Our aim was to study fasting ghrelin and their response to an oral glucose tolerance test (OGTT) in liver failure patients and normal subjects. Methods We included 16 patients with severe liver failure prior to liver transplantation. As a control group we included 10 age- and BMI-matched healthy subjects. After an overnight fast, 75 g of oral glucose were administered; glucose, insulin, and ghrelin were obtained at baseline and at times 30, 60, 90, and 120 min, respectively. Results Fasting ghrelin (median and range) were statistically significantly lower for patients compared to the controls, 527 (377–971) pg/ml vs. 643 (523–2163) pg/ml, P = 0.045, for patients and controls, respectively. The area under the curve for total ghrelin post-OGTT were lower in end-stage liver failure patients than in the control group, 58815 (44730–87420) pg/ml min vs. 76560 (56160–206385) pg/ml min, for patients and controls, respectively, P = 0.027. Conclusions Ghrelin levels are significantly decreased both fasting and post-OGTT in patients with liver failure candidates for transplantation. Decreased ghrelin levels could contribute to anorexia in patients with cirrhosis.Instituto de Salud Carlos III; PI051024Instituto de Salud Carlos III; PI070413Xunta de Galicia; PS07/12Xunta de Galicia; PGIDT05PXIC91605PNXunta de Galicia; INCITE08ENA916110E

Nat Metab

Hypothalamic AgRP and POMC neurons are conventionally viewed as the yin and yang of the body’s energy status, since they act in an opposite manner to modulate appetite and systemic energy metabolism. However, although AgRP neurons’ functions are comparatively well understood, a unifying theory of how POMC neuronal cells operate has remained elusive, probably due to their high level of heterogeneity, which suggests that their physiological roles might be more complex than initially thought. In this Perspective, we propose a conceptual framework that integrates POMC neuronal heterogeneity with appetite regulation, whole-body metabolic physiology and the development of obesity. We highlight emerging evidence indicating that POMC neurons respond to distinct combinations of interoceptive signals and food-related cues to fine-tune divergent metabolic pathways and behaviours necessary for survival. The new framework we propose reflects the high degree of developmental plasticity of this neuronal population and may enable progress towards understanding of both the aetiology and treatment of metabolic disorders.Bordeaux Region Aquitaine Initiative for NeuroscienceInnovations instrumentales et procédurales en psychopathologie expérimentale chez le rongeurLa signalisation des acides biliaires dans le cerveau et son rôle dans le contrôle métaboliqueRôle du récepteur aux cannabinoïdes de type 1 mitochondriale dans les circuits hypothalamiques et son interaction avec la voie mTORC1 dans l'obésité.Rôle de Tbx3 dans la détermination de l'identité fonctionnelle des neurones POMC dans l'obésitéEuropean Union Seventh Framework Programme FP7/2007-201