ABSTRACT
BACKGROUND: Optimizing the refeeding of patients with anorexia nervosa remains important to limit somatic complications of malnutrition, as well as to avoid disease relapses by targeting persistent mood and intestinal disorders. We aimed to evaluate the effects of glutamine (Gln) and branched-chain amino acids (BCAA) supplementation during refeeding in activity-based anorectic (ABA) mice. METHOD: Male C57Bl/6 mice were randomized in control and ABA groups. Once ABA-induced malnutrition was established, mice were progressively refed or not. Refed mice had free access to drinking water supplemented or not with 1% Gln or 2.5% BCAA for 10 days. RESULTS: A progressive refeeding was associated with a partial restoration of body weight and lean mass, while a fat mass rebound was observed. In addition, refeeding restored glucose and leptin. Gln did not affect these parameters, while BCAA tended to increase body weight, fat mass, and glycaemia. In the colon, refeeding improved total protein synthesis and restored the LC3II/LC3I ratio, a marker of autophagy. Gln supplementation enhanced colonic protein synthesis, which was associated with an increased p-p70S6kinase/p70S6kinase ratio, whereas these effects were blunted by BCCA supplementation. CONCLUSIONS: In ABA mice, Gln and BCAA supplementations during a progressive refeeding fail to restore body weight and lean mass. However, Gln supplementation improves total colonic protein synthesis conversely to BCAA. Further studies are needed to decipher the underlying mechanisms involved in these opposite results.
Subject(s)
Amino Acids, Branched-Chain/administration & dosage , Anorexia Nervosa/metabolism , Dietary Supplements , Glutamine/administration & dosage , Malnutrition/metabolism , Animals , Anorexia Nervosa/physiopathology , Body Composition , Colon/physiopathology , Feeding Behavior , Male , Malnutrition/physiopathology , Mice , Mice, Inbred C57BL , Permeability , Protein BiosynthesisABSTRACT
BACKGROUND: During activity-based anorexia (ABA) in mice, enhanced paracellular permeability and reduced protein synthesis have been shown in the colon while the gut-brain axis has received increasing attention in the regulation of intestinal and mood disorders that frequently occur during anorexia nervosa, a severe eating disorder for which there is no specific treatment. In the present study, we assessed the effects of oral glutamine (Gln) or branched-chain amino acids (BCAA) supplementation during ABA to target intestinal functions, body composition and feeding behavior. METHODS: C57BL/6 male mice were randomized in Control (CTRL) and ABA groups. After ABA induction, mice received, or not, either 1% Gln or 2.5% BCAA (Leu, Ile, Val) for one week in drinking water. RESULTS: Neither Gln nor BCAA supplementation affected body weight and body composition, while only Gln supplementation slightly increased food intake. ABA mice exhibited increased paracellular permeability and reduced protein synthesis in the colonic mucosa. Oral Gln restored colonic paracellular permeability and protein synthesis and increased the mucin-2 mRNA level, whereas BCAA did not affect colonic parameters. CONCLUSION: In conclusion, oral Gln specifically improves colonic response during ABA. These data should be further confirmed in AN patients.
Subject(s)
Amino Acids, Branched-Chain/pharmacology , Anorexia/drug therapy , Dietary Supplements , Glutamine/pharmacology , Intestinal Mucosa/drug effects , Animals , Anorexia/physiopathology , Body Composition/drug effects , Colon/drug effects , Colon/physiopathology , Feeding Behavior/drug effects , Male , Mice , Mice, Inbred C57BL , Permeability/drug effects , Protein Biosynthesis/drug effectsABSTRACT
Neurosteroids can modulate the activity of the GABAA receptors, and thus affect anxiety-like behaviors. The non-benzodiazepine anxiolytic compound etifoxine has been shown to increase neurosteroid concentrations in brain tissue but the mode of action of etifoxine on neurosteroid formation has not yet been elucidated. In the present study, we have thus investigated the effect and the mechanism of action of etifoxine on neurosteroid biosynthesis using the frog hypothalamus as an experimental model. Exposure of frog hypothalamic explants to graded concentrations of etifoxine produced a dose-dependent increase in the biosynthesis of 17-hydroxypregnenolone, dehydroepiandrosterone, progesterone and tetrahydroprogesterone, associated with a decrease in the production of dihydroprogesterone. Time-course experiments revealed that a 15-min incubation of hypothalamic explants with etifoxine was sufficient to induce a robust increase in neurosteroid synthesis, suggesting that etifoxine activates steroidogenic enzymes at a post-translational level. Etifoxine-evoked neurosteroid biosynthesis was not affected by the central-type benzodiazepine (CBR) receptor antagonist flumazenil, the translocator protein (TSPO) antagonist PK11195 or the GABAA receptor antagonist bicuculline. In addition, the stimulatory effects of etifoxine and the triakontatetraneuropeptide TTN, a TSPO agonist, were additive, indicating that these two compounds act through distinct mechanisms. Etifoxine also induced a rapid stimulation of neurosteroid biosynthesis from frog hypothalamus homogenates, a preparation in which membrane receptor signalling is disrupted. In conclusion, the present study demonstrates that etifoxine stimulates neurosteroid production through a membrane receptor-independent mechanism.
Subject(s)
17-alpha-Hydroxypregnenolone/agonists , Anti-Anxiety Agents/pharmacology , Dehydroepiandrosterone/agonists , Hypothalamus/drug effects , Oxazines/pharmacology , Pregnanolone/agonists , Progesterone/agonists , 17-alpha-Hydroxypregnenolone/metabolism , 20-alpha-Dihydroprogesterone/antagonists & inhibitors , 20-alpha-Dihydroprogesterone/biosynthesis , Animals , Bicuculline/pharmacology , Complex Mixtures/chemistry , Dehydroepiandrosterone/biosynthesis , Dose-Response Relationship, Drug , Flumazenil/pharmacology , GABA Modulators/pharmacology , GABA-A Receptor Antagonists/pharmacology , Gene Expression , Hypothalamus/metabolism , Isoquinolines/pharmacology , Male , Neuropeptides/pharmacology , Peptide Fragments/pharmacology , Pregnanolone/biosynthesis , Progesterone/biosynthesis , Rana esculenta , Receptors, GABA-A/genetics , Receptors, GABA-A/metabolism , Tissue Culture TechniquesABSTRACT
The sulfated neurosteroids pregnenolone sulfate (Δ(5)PS) and dehydroepiandrosterone sulfate (DHEAS) are known to play a role in the control of reproductive behavior. In the frog Pelophylax ridibundus, the enzyme hydroxysteroid sulfotransferase (HST), responsible for the biosynthesis of Δ(5)PS and DHEAS, is expressed in the magnocellular nucleus and the anterior preoptic area, two hypothalamic regions that are richly innervated by GnRH1-containing fibers. This observation suggests that GnRH1 may regulate the formation of sulfated neurosteroids to control sexual activity. Double labeling of frog brain slices with HST and GnRH1 antibodies revealed that GnRH1-immunoreactive fibers are located in close vicinity of HST-positive neurons. The cDNAs encoding 3 GnRH receptors (designated riGnRHR-1, -2, and -3) were cloned from the frog brain. RT-PCR analyses revealed that riGnRHR-1 is strongly expressed in the hypothalamus and the pituitary whereas riGnRHR-2 and -3 are primarily expressed in the brain. In situ hybridization histochemistry indicated that GnRHR-1 and GnRHR-3 mRNAs are particularly abundant in preoptic area and magnocellular nucleus whereas the concentration of GnRHR-2 mRNA in these 2 nuclei is much lower. Pulse-chase experiments using tritiated Δ(5)P and DHEA as steroid precursors, and 3'-phosphoadenosine 5'-phosphosulfate as a sulfonate moiety donor, showed that GnRH1 stimulates, in a dose-dependent manner, the biosynthesis of Δ(5)PS and DHEAS in frog diencephalic explants. Because Δ(5)PS and DHEAS, like GnRH, stimulate sexual activity, our data strongly suggest that some of the behavioral effects of GnRH could be mediated via the modulation of sulfated neurosteroid production.
Subject(s)
Dehydroepiandrosterone Sulfate/metabolism , Gonadotropin-Releasing Hormone/metabolism , Hypothalamus/metabolism , Pregnenolone/metabolism , Amino Acid Sequence , Animals , Cell Line , Diencephalon/drug effects , Diencephalon/metabolism , Gene Expression Profiling , Gonadotropin-Releasing Hormone/pharmacology , In Situ Hybridization , Male , Microscopy, Confocal , Molecular Sequence Data , Neurons/metabolism , Pituitary Gland/metabolism , Protein Isoforms/genetics , Protein Isoforms/metabolism , Ranidae , Receptors, LHRH/genetics , Receptors, LHRH/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Sulfotransferases/metabolismABSTRACT
Hypothalamic glucose sensing is involved in the control of feeding behavior and peripheral glucose homeostasis, and glial cells are suggested to play an important role in this process. Diazepam-binding inhibitor (DBI) and its processing product the octadecaneuropeptide (ODN), collectively named endozepines, are secreted by astroglia, and ODN is a potent anorexigenic factor. Therefore, we investigated the involvement of endozepines in brain glucose sensing. First, we showed that intracerebroventricular administration of glucose in rats increases DBI expression in hypothalamic glial-like tanycytes. We then demonstrated that glucose stimulates endozepine secretion from hypothalamic explants. Feeding experiments indicate that the anorexigenic effect of central administration of glucose was blunted by coinjection of an ODN antagonist. Conversely, the hyperphagic response elicited by central glucoprivation was suppressed by an ODN agonist. The anorexigenic effects of centrally injected glucose or ODN agonist were suppressed by blockade of the melanocortin-3/4 receptors, suggesting that glucose sensing involves endozepinergic control of the melanocortin pathway. Finally, we found that brain endozepines modulate blood glucose levels, suggesting their involvement in a feedback loop controlling whole-body glucose homeostasis. Collectively, these data indicate that endozepines are a critical relay in brain glucose sensing and potentially new targets in treatment of metabolic disorders.
Subject(s)
Appetite Regulation , Diazepam Binding Inhibitor/metabolism , Feedback, Physiological , Glucose/metabolism , Hypothalamus/metabolism , Neuroglia/metabolism , Neuropeptides/metabolism , Peptide Fragments/metabolism , Animals , Appetite Depressants/administration & dosage , Appetite Depressants/pharmacology , Appetite Regulation/drug effects , Appetite Stimulants/administration & dosage , Appetite Stimulants/pharmacology , Appetitive Behavior/drug effects , Arcuate Nucleus of Hypothalamus/cytology , Arcuate Nucleus of Hypothalamus/drug effects , Arcuate Nucleus of Hypothalamus/metabolism , Diazepam Binding Inhibitor/agonists , Diazepam Binding Inhibitor/antagonists & inhibitors , Feedback, Physiological/drug effects , Gene Expression Regulation/drug effects , Glucose/administration & dosage , Hypothalamus/cytology , Hypothalamus/drug effects , Injections, Intraventricular , Male , Nerve Tissue Proteins/agonists , Nerve Tissue Proteins/antagonists & inhibitors , Nerve Tissue Proteins/metabolism , Neuroglia/drug effects , Neuropeptides/antagonists & inhibitors , Peptide Fragments/antagonists & inhibitors , Protein Processing, Post-Translational , Rats , Rats, Wistar , Receptors, Melanocortin/antagonists & inhibitors , Receptors, Melanocortin/metabolism , Synaptic Transmission/drug effects , Tissue Culture TechniquesABSTRACT
We have recently shown that hydroxysteroid sulfotransferase (HST), the enzyme responsible for the biosynthesis of pregnenolone sulfate (Delta(5)PS) and dehydroepiandrosterone sulfate (DHEAS), is expressed in neurons located in the anterior preoptic area and the dorsal magnocellular nucleus of the frog diencephalon. As these two nuclei are richly innervated by NPY-immunoreactive fibers, we investigated the possible implication of NPY in the control of Delta(5)PS and DHEAS biosynthesis. Double labeling of frog brain sections revealed that 42% of the HST-immunoreactive perikarya in the diencephalon were contacted by NPY-containing fibers. In situ hybridization studies showed that Y(1) and Y(5) receptor mRNAs are expressed in the anterior preoptic area and the dorsal magnocellular nucleus. Pulse-chase experiments with (35)S-labeled 3'-phosphoadenosine 5'-phosphosulfate as a sulfate donor demonstrated that frog NPY (fNPY) inhibited the conversion of [(3)H]Delta(5)P and [(3)H]dehydroepiandrosterone ([(3)H]DHEA) into [(3)H,(35)S]Delta(5)PS and [(3)H,(35)S]DHEAS by diencephalic explants. The inhibitory effect of fNPY on Delta(5)PS and DHEAS formation was mimicked by (pPYY) and [Leu(31),Pro(34)]pNPY, which is an agonist for non-Y(2) receptors in mammals, and was completely suppressed by the Y(1) receptor antagonist BIBP3226. Conversely, the Y(2) receptor agonist pNPY-(13-36) and the Y(5) receptor agonist [D-Trp(32)]pNPY did not significantly modify the biosynthesis of [(3)H,(35)S]Delta(5)PS and [(3)H,(35)S]DHEAS. The present study provides the first evidence for the innervation of neurosteroid-producing neurons by NPY fibers. Our data also demonstrate that NPY, acting via Y(1) receptors, exerts an inhibitory effect on the biosynthesis of sulfated neurosteroids.