Hypophagia induced by salmon calcitonin, but not by amylin, is partially driven by malaise and is mediated by CGRP neurons

Objective: The behavioral mechanisms and the neuronal pathways mediated by amylin and its long-acting analog sCT (salmon calcitonin) are not fully understood and it is unclear to what extent sCT and amylin engage overlapping or distinct neuronal subpopulations to reduce food intake. We here hypothesize that amylin and sCT recruit different neuronal population to mediate their anorectic effects. Methods: Viral approaches were used to inhibit calcitonin gene-related peptide (CGRP) lateral parabrachial nucleus (LPBN) neurons and assess their role in amylin's and sCT's ability to decrease food intake in mice. In addition, to test the involvement of LPBN CGRP neuropeptidergic signaling in the mediation of amylin and sCT's effects, a LPBN site-specific knockdown was performed in rats. To deeper investigate whether the greater anorectic effect of sCT compared to amylin is due do the recruitment of additional neuronal pathways related to malaise multiple and distinct animal models tested whether amylin and sCT induce conditioned avoidance, nausea, emesis, and conditioned affective taste aversion. Results: Our results indicate that permanent or transient inhibition of CGRP neurons in LPBN blunts sCT-, but not amylin-induced anorexia and neuronal activation. Importantly, sCT but not amylin induces behaviors indicative of malaise including conditioned affective aversion, nausea, emesis, and conditioned avoidance; the latter mediated by CGRPLPBN neurons. Conclusions: Together, the present study highlights that although amylin and sCT comparably decrease food intake, sCT is distinctive from amylin in the activation of anorectic neuronal pathways associated with malaise

Neuronal mechanisms involved in cancer anorexia-cachexia syndrome and evaluation of possible therapeutic approaches

The cancer anorexia-cachexia syndrome (CACS) is characterized by reduced eating and body weight loss, which is the consequence of reduced fat and muscle mass. CACS is present in up to 85% of cancer patients and represents the most significant negative predictor of treatment outcome. Importantly, it also deteriorates quality of life and leads to severe psychological suffering of patients and their families. The currently available therapeutic approaches are not sufficiently effective and poorly understood as far as the underlying mode of action is concerned. Therefore, the identification of the underlying pathophysiology and the development of new pharmacological approaches against CACS have a high clinical relevance. The area postrema (AP) and the nucleus tractus solitarii (NTS) are two important brainstem centers implicated in the mediation of anorexia, emesis and nausea during sickness conditions. However, the knowledge about the role of the AP and NTS in CACS is very limited. Recently evidence suggests that the macrophage inhibitory cytokine-1 (MIC-1) acts as circulating tumor-derived factor promoting anorexia and body weight loss. Using neurosurgical approaches we examined the role of the AP and vagal afferents projecting to the NTS in the mediation of cancer anorexia and body weight loss in a rat hepatoma tumor model. Specifically, we investigated whether a lesion of the AP or subdiaphragmatic vagal deafferentation attenuate anorexia, body weight, muscle and fat loss. Moreover, MIC-1 blood levels were analyzed at different time points during cancer progression. AP lesioned rats were protected against anorexia, body weight loss and muscle atrophy induced by tumor growth. In contrast, the subdiaphragmatic vagal deafferentation did not attenuate cancer-induced anorexia or bodyweight loss. Tumor-bearing (TB) rats had substantially increased MIC-1 levels, which positively correlated with tumor size and cancer progression, and negatively correlated with food intake. These findings demonstrate the importance of the AP in the central mediation of CACS in our tumor model and support a pathological role of MIC-1 as a tumor-derived factor mediating CACS, possibly via an AP-dependent action. The glucagon-like peptide-1 (GLP-1) is expressed in neurons of the NTS. GLP-1ergic neurons project from the NTS to the AP where the GLP-1 receptor (GLP-1R) is highly expressed. GLP-1 is implicated in malaise, nausea and aversion induced by different stimuli. Therefore, we hypothesized that GLP-1 signaling in the AP/NTS region also mediates anorexia following tumor growth. We first tested whether a pharmacological blockade of brainstem GLP-1 via chronic delivery of the GLP-1R antagonist exendin-9 (Ex-9) into the fourth ventricle attenuated CACS. Second, we investigated whether a genetic knockdown of GLP-1 expression in the NTS ameliorates CACS. Finally, using a two diets choice paradigm, we tested if tumor growth in our model is paralleled by the development of food aversion. Pharmacological or genetic blockade of brainstem GLP-1 signaling attenuated anorexia, body weight loss and fat depletion. Importantly, Ex-9 treatment was also effective in preventing muscle degradation. Furthermore, TB rats developed tumor-induced food aversion because their flavor preference turned into a flavor rejection during tumor growth. Our study identifies brainstem GLP-1 as crucial mediator of CACS in hepatoma TB rats. In line with the function of GLP-1 as a mediator of aversion, emesis and nausea, we also demonstrated the development of food aversion during CACS in this model. Since the AP lacks a functional blood-brain-barrier, the GLP-1R located in the AP represents a promising target for pharmacological approaches against CACS. In addition to brainstem mechanisms, forebrain structures in particular the arcuate nucleus (Arc) seem to be implicated in the mediation of CACS. Hence, the underlying signaling cascades influencing Arc activity represent potential therapeutic targets. The gastric hormone ghrelin positively affects energy balance by increasing food intake and reducing energy expenditure. These effects are mainly mediated via the Arc. Therefore, ghrelin mimetics are another possible treatment against CACS. Using the aforementioned rat hepatoma model, the action of the non-peptidergic ghrelin receptor agonist HM01 energy homeostasis and muscle mass was evaluated. TB rats treated with HM01 had higher food intake than TB controls and were protected against body weight loss and muscle degradation. Moreover, TB rats reduced their metabolic rate following HM01 treatment. These results emphasize the possible therapeutic usefulness of ghrelin receptor agonists like HM01 for the treatment of CACS. In conclusion, we substantiated the fundamental role of the AP in the mediation of CACS in our tumor model and we provided evidence for a role of central GLP-1 signaling in hepatoma TB rats. Furthermore, we demonstrated the therapeutic efficacy of the novel ghrelin analog HM01 in the context of CACS. Both ghrelin and GLP-1-based approaches represent promising options for the treatment of CACS and possibly other forms of disease-related anorexia. Das krebsbedingte Anorexie-Kachexie-Syndrom (engl. CACS) zeichnet sich durch einen Verlust des Körpergewichts aus, welchem eine verminderte Nahrungsaufnahme und ein erhöhter Fett- und Muskelmasseabbau zugrunde liegt. CACS tritt bei bis zu 85% der Krebspatienten auf und ist der wichtigste Faktor, welcher den Therapieerfolg negativ beeinflusst. Zusätzlich reduziert CACS die Lebensqualität und kann schwere psychische Probleme bei Patienten und deren Familien hervorrufen. Leider sind die derzeit verfügbaren therapeutischen Ansätze nicht wirksam genug und deren Mechanismen unbekannt. Folglich ist die Erforschung der zugrundeliegenden Pathophysiologie sowie die Entwicklung neuer pharmakologischer Ansätze von hoher klinischer Bedeutung. Die Area postrema (AP) und der Nucleus tractus solitarii (NTS) sind zwei wichtige Zentren im Hirnstamm, die bei der physiologischen Kontrolle der Nahrungsaufnahme und der Vermittlung krankheitsbedingter Anorexie sowie bei Übelkeit und Erbrechen beteiligt sind. Ihre Beteiligung bei der Entstehung von CACS ist derzeit noch unklar. Als ein wichtiger tumor-assoziierter Faktor wurde kürzlich das Macrophage inhibitory cytokine-1 (MIC-1) identifiziert, welches Anorexie und Körpergewichtsverlust, wahrscheinlich durch einen direkten zentralen Effekt auslösen kann. Mittels neurochirurgischer Ansätze haben wir die Rolle der AP und der vagalen Afferenzen, die zum NTS projizieren, bei der Vermittlung von CACS anhand eines Ratten Hepatoma- Tumormodells untersucht. Dabei haben wir erforscht, ob eine Läsion der AP oder die Deafferenzierung der subdiaphragmatischen vagalen Afferenzen der krebsbedingten Reduktion von Futteraufnahme, Körpergewicht sowie Muskel- und Fettmasse entgegenwirkt. Ausserdem haben wir die Plasmakonzentration von MIC-1 zu verschiedenen Zeitpunkten während des Tumorwachstums untersucht. Im Gegensatz zu der subdiaphragmatischen vagalen Deafferenzierung hob eine Läsion der AP die tumorbedingte Anorexie und den Körpergewichtsverlust der Ratten auf. Die MIC-1 Plasmakonzentration war bei tumortragenden Ratten signifikant erhöht und korrelierte positiv mit der Tumorgrösse und negativ mit der Nahrungsaufnahme. Diese Ergebnisse zeigen die Wichtigkeit der AP bei der Entwicklung der krebsbedingten Anorexie und Körpergewichtsreduktion in unserem Tumormodell und stützen die Annahme, dass MIC-1 eine wichtige Rolle bei der Entwicklung von CACS spielt und vermutlich über die AP wirkt. Das Peptid „Glucagon-like peptide-1“ (GLP-1) wird unter anderem in Neuronen des NTS exprimiert. Diese Neuronen projizieren zur AP, die eine hohe Expression des GLP-1 Rezeptor (GLP-1R) aufweist. GLP-1 spielt eine wichtige Rolle bei der Entstehung von Unwohlsein, Übelkeit und Futteraversion, die durch verschiedene Reize ausgelöst werden können. Wir vermuteten, dass auch die tumorbedingte Anorexie über GLP-1 in der AP/NTS-Region vermittelt werden könnte. Zunächst haben wir untersucht, wie sich eine pharmakologische Blockade vom GLP-1R im Hirnstamm durch Infusion des GLP- 1R-Antagonisten Exendin-9 (Ex-9) in den vierten Ventrikel auf das CACS auswirkt. Zusätzlich haben wir getestet, ob eine Reduktion der GLP-1 Genexpression im NTS von Ratten durch einen genetischen GLP-1 „knockdown“ vor Anorexie und Körpergewichtsverlust bei Tumoren schützt. Schlussendlich haben wir getestet, ob das Tumorwachstum in unserem Modell mit der Entstehung einer Nahrungsaversion assoziiert ist. Die pharmakologische sowie auch die genetische Inhibition von GLP-1 im Hirnstamm führte zu verminderter Anorexie und reduziertem Körpergewichts- und Fettverlust. Die Ex-9 Behandlung verminderte zusätzlich den Muskelabbau. Ausserdem entwickelten tumortragende Ratten eine tumorinduzierte Nahrungsaversion gegen das Futter, das sie vor Beginn des Tumorwachstums besonders gern zu sich nahmen. Unsere Studien zeigen somit, dass GLP-1 im Hirnstamm eine entscheidende Rolle bei der Entwicklung von CACS in tumortragenden Ratten spielt. Zusätzlich haben wir in diesem Modell ebenfalls die Entwicklung einer Nahrungsaversion im Zusammenhang mit CACS gezeigt. Da der AP eine funktionelle Blut-Hirn-Schranke fehlt, sind die GLP-1 Rezeptoren in der AP ein möglicher Angriffspunkt für neuartige Therapien gegen CACS. Zusätzlich zu den Hirnstammmechanismen sind offensichtlich Strukturen im Vorderhirn, insbesondere der Nucleus Arcuatus (Arc), an der Vermittlung des CACS beteiligt. Daher stellt auch der Arc ein potenzielles therapeutisches Ziel dar. Das Hormon Ghrelin wirkt sich positiv auf die Energiebilanz aus, indem es die Nahrungsaufnahme erhöht und den Energieverbrauch senkt. Diese Effekte werden hauptsächlich über den Arc vermittelt. Deswegen stellen Ghrelin-Mimetika grundsätzlich eine weitere Behandlungsmöglichkeit gegen CACS dar. Mittels unseres Tumormodells in Ratten untersuchten wir die Wirkung des Ghrelin-Rezeptor-Agonisten HM01 auf die Energiehomöostase und die Muskelmasse bei Ratten. Tumortragende Ratten, die mit HM01 behandelt wurden, hatten eine höhere Futteraufnahme als die tumortragenden Kontrollen sowie einen geringeren Körpergewichtsverlust und Muskelabbau. Außerdem reduzierten die mit HM01 behandelten Ratten ihre Stoffwechselrate. Diese Ergebnisse legen eine mögliche therapeutische Nutzung von Ghrelin-Rezeptor-Agonisten wie HM01 für die Behandlung von CACS nahe. Zusammenfassend haben wir die Bedeutung der AP und des NTS bei der Vermittlung des CACS in unserem Tumormodell untersucht und haben dabei GLP-1 als massgeblich für die Signalvermittlung identifiziert. Weiterhin haben wir die therapeutische Wirksamkeit des Ghrelin-Analogs HM01 bei CACS nachgewiesen. Sowohl Ghrelin- als auch GLP-1-basierte therapeutische Ansätze stellen vielversprechende Möglichkeiten für die Behandlung von CACS und eventuell auch für andere Formen von krankheitsbedingter Anorexie dar

Lipopolysaccharide inhibits ghrelin-excited neurons of the arcuate nucleus and reduces food intake via central nitric oxide signaling

Lipopolysaccharide (LPS) induces anorexia and expression of inducible nitric oxide synthase (iNOS) in the hypothalamic arcuate nucleus (Arc). Peripheral administration of the iNOS inhibitor 1400W counteracts the anorectic effects of LPS. Here we investigated the role of central NO signaling in LPS anorexia. In electrophysiological studies we tested whether 1400W counteracts the iNOS-dependent inhibition of Arc neurons triggered by in vivo or in vitro stimulation with LPS. We used the hormone ghrelin as a functional reference stimulus because ghrelin is known to activate orexigenic Arc neurons. Further, we investigated whether in vitro LPS stimulation induces an iNOS-mediated formation of the second messenger cGMP. Since the STAT1 pathway contributes to the regulation of iNOS expression we investigated whether LPS treatment induces STAT1 phosphorylation in the Arc. Finally we tested the effect of intracerebroventricular injection of 1400W on LPS-induced anorexia. Superfusion with 1400W (10(-4)M) increased neuronal activity in 37% of neurons in Arc slices from LPS treated (100μg/kg ip) but not from saline treated rats. Similarly, 1400W excited 45% of Arc neurons after in vitro stimulation with LPS (100ng/ml). In both approaches, a considerable percentage of 1400W sensitive neurons were excited by ghrelin (10(-8)M; 50% and 75%, respectively). In vitro stimulation with LPS induced cGMP formation in the Arc, which was blocked by co-incubation with 1400W. LPS treatment elicited a pSTAT1 response in the Arc of mice. Central 1400W injection (4μg/rat) attenuated LPS-induced anorexia and counteracted the LPS-dependent decrease in respiratory quotient and energy expenditure. In conclusion, the current findings substantiate a role of central iNOS dependent NO formation in LPS-induced effects on eating and energy homeostasis. A pharmacological blockade of NO formation might be a therapeutic approach to ameliorate disease-related anorexia

Brainstem GLP-1 signalling contributes to cancer anorexia-cachexia syndrome in the rat

The cancer anorexia-cachexia syndrome (CACS) is a frequent and severe condition in cancer patients. Currently, no pharmacological treatment is approved for the therapy of CACS. Centrally, glucagon-like peptide-1 (GLP-1) is expressed in the nucleus tractus solitarii (NTS) and is implicated in malaise, nausea and food aversion. The NTS is reciprocally connected to brain sites implicated in the control of energy balance including the area postrema (AP), which mediates CACS in certain tumour models. Given the role of GLP-1 as a mediator of anorexia under acute sickness conditions, we hypothesized that brainstem GLP-1 signalling might play a role in the mediation of CACS. Using hepatoma tumour-bearing (TB) rats, we first tested whether the chronic delivery of the GLP-1R antagonist exendin-9 (Ex-9) into the fourth ventricle attenuates CACS. Second, we investigated whether a genetic knockdown of GLP-1 expression in the NTS ameliorates CACS. Ex-9 attenuated anorexia, body weight loss, muscle and fat depletion compared to TB controls. Similarly, TB animals with a knockdown of GLP-1 expression in the NTS had higher food intake, reduced body weight loss, and higher lean and fat mass compared to TB controls. Our study identifies brainstem GLP-1 as crucial mediator of CACS in hepatoma TB rats. The GLP-1R represents a promising target against CACS and possibly other forms of disease-related anorexia/cachexia

The satiating hormone amylin enhances neurogenesis in the area postrema of adult rats

Objective: Adult neurogenesis in the subgranular zone and subventricular zone is generally accepted, but its existence in other brain areas is still controversial. Circumventricular organs, such as the area postrema (AP) have recently been described as potential neurogenic niches in the adult brain. The AP is the major site of action of the satiating hormone amylin. Amylin has been shown to promote the formation of neuronal projections originating from the AP in neonatal rodents but the role of amylin in adult neurogenesis remains unknown. Methods: To test this, we first performed an RNA-sequencing of the AP of adult rats acutely injected with either amylin (20 μg/kg), amylin plus the amylin receptor antagonist AC187 (500 μg/kg) or vehicle. Second, animals were subcutaneously equipped with minipumps releasing either amylin (50 μg/kg/day) or vehicle for 3 weeks to assess cell proliferation and differentiation with the 5′-bromo-2-deoxyuridine (BrdU) technique. Results: Acute amylin injections affected genes involved in pathways and processes that control adult neurogenesis. Amylin consistently upregulated NeuroD1 transcript and protein in the adult AP, and this effect was blocked by the co-administration of AC187. Further, chronic amylin treatment increased the number of newly proliferated AP-cells and significantly promoted their differentiation into neurons rather than astrocytes. Conclusion: Our findings revealed a novel role of the satiating hormone amylin in promoting neurogenesis in the AP of adult rats. Keywords: Amylin, Adult neurogenesis, Area postrema, BrdU, Circumventricular organ

Direct conservation payments in the Brazilian Amazon: Scope and equity implications

This article looks into the scope and equity implications of applying payments for environmental services (PES) as a REDD implementation mechanism in the Brazilian Amazon. We establish a set of economic and institutional preconditions for PES to become a feasible and cost-effective conservation mechanism. We proceed with a macro-scale spatial analysis and overlay of opportunity costs, deforestation patterns, carbon services, and land tenure, in order to assess where these conditions hold. We then screen how the benefits of potential PES schemes might be distributed across different socioeconomic groups of service providers in different land tenure categories. Our economic–quantitative analysis, though sensitive to documented assumptions, suggests that under current carbon prices the economic preconditions are in place to pay for avoided deforestation in over half of threatened forests over the next decade. Unfortunately, the same optimism does not apply to institutional preconditions. Land grabbing, insecure tenure, overlapping claims, and lacking information on private tenure constitute real medium-term impediments to PES. If payments were to accrue to current landholders regardless of current tenure insecurities, large landowners who account for about 80% of all deforestation would reap the highest benefits, though per-capita benefits other tenure categories are also high. Schemes that closely align payments with opportunity costs are preferable for cost-effectiveness, and not necessarily more inequitable in outcomes. Essentially, PES systems cannot substitute command-and-control measures: the former depend on the latter for basic governance systems to secure effective rights of exclusion, which land stewards essentially need in order to become reliable service providers

The ghrelin receptor agonist HM01 mimics the neuronal effects of ghrelin in the arcuate nucleus and attenuates anorexia-cachexia syndrome in tumor-bearing rats

The gastric hormone ghrelin positively affects energy balance by increasing food intake and reducing energy expenditure. Ghrelin mimetics are a possible treatment against cancer anorexia-cachexia syndrome (CACS). This study aimed to characterize the action of the nonpeptidergic ghrelin receptor agonist HM01 on neuronal function, energy homeostasis and muscle mass in healthy rats and to evaluate its possible usefulness for the treatment of CACS in a rat tumor model. Using extracellular single-unit recordings, we tested whether HM01 mimics the effects of ghrelin on neuronal activity in the arcuate nucleus (Arc). Furthermore, we assessed the effect of chronic HM01 treatment on food intake (FI), body weight (BW), lean and fat volumes, and muscle mass in healthy rats. Using a hepatoma model, we investigated the possible beneficial effects of HM01 on tumor-induced anorexia, BW loss, muscle wasting, and metabolic rate. HM01 (10(-7)-10(-6) M) mimicked the effect of ghrelin (10(-8) M) by increasing the firing rate in 76% of Arc neurons. HM01 delivered chronically for 12 days via osmotic minipumps (50 μg/h) increased FI in healthy rats by 24%, paralleled by increased BW, higher fat and lean volumes, and higher muscle mass. Tumor-bearing rats treated with HM01 had 30% higher FI than tumor-bearing controls and were protected against BW loss. HM01 treatment resulted in higher muscle mass and fat mass. Moreover, tumor-bearing rats reduced their metabolic rate following HM01 treatment. Our studies substantiate the possible therapeutic usefulness of ghrelin receptor agonists like HM01 for the treatment of CACS and possibly other forms of disease-related anorexia and cachexia