The Anti‐Ghrelin Spiegelmer NOX‐B11‐3 Blocks Ghrelin‐ but not Fasting‐Induced Neuronal Activation in the Hypothalamic Arcuate Nucleus

Abstract

The anti-ghrelin Spiegelmer NOX-B11-3 blocks ghrelin-but not fasting-induced neuronal activation in the hypothalamic arcuate nucleus AbstractThe hypothalamic arcuate nucleus (Arc) is the presumed target site for the orexigenic hormone ghrelin which is secreted from the stomach during fasting. Ghrelin directly activates Arc neurones. Similar to exogenous ghrelin, overnight food deprivation also induces c-Fos expression in the Arc of mice. In this study we tested the role of endogenous ghrelin in the fasting-induced c-Fos expression in the Arc of mice. We used NOX-B11-3, the latest generation of the recently developed ghrelin-binding compounds, so-called RNA Spiegelmers (SPM) to block endogenous ghrelin action during food deprivation. The specificity and potency of this compound was also tested in electrophysiological and immunohistological experiments. In electrophysiological in vitro single cell recordings NOX-B11-3 effectively blocked the excitatory effect of ghrelin in the medial Arc (ArcM) of rats while the biologically inactive control SPM had no effect. Furthermore, NOX-B11-3 (15 mg/kg ip) potently suppressed ghrelin-induced (25 µg/kg sc, 12h after SPM injection) c-Fos expression in the Arc. However, the same dose of NOX-B11-3 when injected at the beginning of a 14h fasting period had no effect on the c-Fos expression in the Arc of mice. These results demonstrate that NOX-B11-3 is a long acting compound, which effectively blocks the effect of exogenous ghrelin on neuronal activity in the Arc under in vitro and in vivo conditions. Furthermore, increased ghrelin signalling does not appear to be a necessary factor for the activation of Arc neurones during food deprivation or other fasting-related signals might have masked or compensated the loss of the ghrelin effect.The anti-ghrelin Spiegelmer NOX-B11-3 blocks ghrelin-but not fasting-induced neuronal activation in the hypothalamic arcuate nucleus AbstractThe hypothalamic arcuate nucleus (Arc) is the presumed target site for the orexigenic hormone ghrelin which is secreted from the stomach during fasting. Ghrelin directly activates Arc neurones. Similar to exogenous ghrelin, overnight food deprivation also induces c-Fos expression in the Arc of mice. In this study we tested the role of endogenous ghrelin in the fasting-induced c-Fos expression in the Arc of mice. We used NOX-B11-3, the latest generation of the recently developed ghrelin-binding compounds, so-called RNA Spiegelmers (SPM) to block endogenous ghrelin action during food deprivation. The specificity and potency of this compound was also tested in electrophysiological and immunohistological experiments. In electrophysiological in vitro single cell recordings NOX-B11-3 effectively blocked the excitatory effect of ghrelin in the medial Arc (ArcM) of rats while the biologically inactive control SPM had no effect. Furthermore, NOX-B11-3 (15 mg/kg ip) potently suppressed ghrelin-induced (25 µg/kg sc, 12h after SPM injection) c-Fos expression in the Arc. However, the same dose of NOX-B11-3 when injected ...

Anti-ghrelin Spiegelmer inhibits exogenous ghrelin-induced increases in food intake, hoarding, and neural activation, but not food deprivation-induced increases

“…None of these responses were blocked by SPM in food-deprived hamsters. This inability of SPM to stem these food deprivation-induced increases in ingestive behavior and neural activation, however, is consistent with its inability to prevent food deprivation-induced increases in food intake in laboratory rats (58) and c-Fos-ir in laboratory mice (8). We assayed acylated (active) and des-acyl (inactive) ghrelin in fed and food-deprived SPM-treated hamsters and found large, increases in acyl ghrelin and des-acyl (inactive) ghrelin.…”

“…The subsequent changes in these ingestive behaviors and c-Fos immunoreactivity (c-Fos-ir), an indicator of neural activation (33), and acyl/des-acyl ghrelin were assayed. Changes in cFos-ir in this and other studies of ghrelin and SPM (8,10,43) indicate potential neural activation sites associated with alterations in ingestive behaviors due to ghrelin or lack of ghrelin.…”

“…In two previous studies (8,59), the inability of SPM to block food deprivation-induced increases in food intake and c-Fos-ir was speculated to be due to, among other possibilities, ghrelin not being necessary for these responses. This overlooked another possibility suggested by the increased ghrelin concentrations assayed (58,59)-that the ghrelin assay recognized acylated ghrelin bound to SPM and/or unbound SPM.…”

“…Representative photographs of ARC c-Fos-ir are shown (Fig. 2B) at approximately the same coronal level of the Arc as depicted for similarly SPMtreated mice (8).…”

“…The three groups were 1) saline ϩ 30 g/kg ghrelin, 2) 18 mg/kg SPM ϩ saline, and 3) 18 mg/kg SPM ϩ 30 g/kg ghrelin (SPM was generously provided by NOXXON Pharma AG, Berlin, Germany). The ghrelin dose was selected on the basis of our previous study, showing this dose produced circulating acylated ghrelin concentrations equivalent to that for 48-h food deprivation in Siberian hamsters (39), whereas the SPM dose was calculated by extrapolating from previous studies in laboratory rats and mice (8,43) and our own pilot study in Siberian hamsters (data not shown). A saline ϩ saline group was not included because pilot data showed no difference between saline ϩ saline-and SPM ϩ saline-treated animals for any measure (data not shown).…”