Silvia Q. Giraudo

Feeding effects of hypothalamic injection of melanocortin 4 receptor ligands

It has been reported that intraventricular administration of the melanocortin 4 receptor (MC4-R) agonist MT II and antagonist SHU9119 alter food intake. We found that MT II and SHU9119 have extremely potent effects on feeding when injected in the paraventricular nucleus (PVN), a site where MC4-R gene expression is very high. Our finding provides direct evidence that MC4-R signaling is important in mediating food intake and that melanocortin neurons in the PVN exert a tonic inhibition of feeding behavior. Chronic disruption of this inhibitory signal is a possible explanation of the agouti-obesity syndrome.

Paraventricular hypothalamic α-melanocyte-stimulating hormone and MTII reduce feeding without causing aversive effects

alpha-Melanocyte-stimulating hormone (alpha-MSH) appears to play a tonic inhibitory role in feeding and energy storage. MTII, a specific synthetic MC3-R/MC4-R agonist, has similar effects on feeding in rats. The current studies demonstrate that PVN administration of alpha-MSH or MTII decreases nocturnal and NPY-stimulated food intake without causing aversive effects. Co-administration with NPY of 600 pmol alpha-MSH or 1 pmol MTII into the PVN caused a significant decrease in NPY-induced feeding. PVN administration of MTII or alpha-MSH at doses effective to suppress feeding did not cause conditioned taste aversion (CTA). ICV administration of alpha-MSH, however, did cause weak CTA. These results indicate that the potent effects on feeding of MC3-R and MC4-R agonists when injected into the PVN are not due to aversive effects.

Effects of the opioid antagonist naltrexone on feeding induced by DAMGO in the central nucleus of the amygdala and in the paraventricular nucleus in the rat

The paraventricular nucleus of the hypothalamus (PVN) and the central nucleus of the amygdala (CNA) are two forebrain structures which are important in regulation of ingestive behavior. DAMGO is one of the most reliable and potent mu-selective opioid ligands that increases feeding in both of these brain nuclei. Administration of naloxone, an opioid antagonist, into the CNA prior to DAMGO blocks DAMGO-induced increases in food intake. The effect of this drug combination on food intake has not been evaluated in the PVN. However, intra-PVN injection of naloxone decreases deprivation and NPY-induced feeding. It has been suggested that CNA may modulate activity of midbrain and caudal brainstem centers via the hypothalamus. Based on these data, we evaluated whether an opioid-opioid interaction is present between the CNA and PVN which might affect feeding behavior. To test this, rats were doubly cannulated with 1 cannula placed in the PVN and 1 cannula in the CNA, allowing for co-administration of the opioid agonist into the PVN and the opioid antagonist into the CNA, and vice versa. CNA DAMGO increased feeding more than two-fold as compared to the vehicle-injected rats. When doses of 10, 12.5 and 25 micrograms of naltrexone (NTX) were injected into the PVN, CNA DAMGO no longer increased food intake above control levels. In the reverse situation, PVN DAMGO also increased food intake above control levels. However, when NTX was administrated unilaterally into the CNA at a relatively high dose (25 micrograms) or bilaterally (12.5 micrograms), PVN DAMGO-induced feeding was not altered. This suggests that an opioid-opioid signaling pathway exists from the CNA to the PVN which influences feeding via mu opioid receptors, whereas such a pathway from the PVN to the CNA does not seem to exist.

Association between the amygdala and nucleus of the solitary tract in μ-opioid induced feeding in the rat

The central nucleus of the amygdala (CNA) and the nucleus of the solitary tract (NTS) are important in the regulation of ingestive behavior. We evaluated whether opioid-opioid signaling between the CNA and rostral NTS (rNTS) affect feeding behavior. To test this, rats were doubly cannulated with one cannula placed in the rNTS and one cannula in the CNA, allowing for co-administration of an opioid agonist into one site and an opioid antagonist into the other. Tyr-D-Ala-Gly-(me) Phe-Gly-ol (DAMGO) (2 nmol) injected into the CNA (CNA DAMGO) increased feeding more than two-fold compared to the vehicle-injected rats. This increase in food intake was blocked when doses of 26.5 and 79 nmol of naltrexone (NTX) were injected into the rNTs. In the reverse situation, rNTS DAMGO increased food intake above control levels, and CNA NTX blocked DAMGO-induced feeding when administrated in doses of 26.5 and 79 nmol. This suggests that a bi-directional opioid-opioid signaling pathway exists between the CNA and the rNTS which influences feeding via mu-opioid receptors.

Agouti-related protein in the hypothalamic paraventricular nucleus: effect on feeding.

Agouti-related protein (Agrp) is an endogenous melanocortin-4 receptor antagonist implicated in the regulation of food intake. Effects of Agrp on feeding under varying conditions were investigated. Agrp (10 to 100 pmol) was injected into the hypothalamic paraventricular nucleus of satiated (a.m. and p.m. injections) and food-deprived rats, or was co-administered with 117 pmol Neuropeptide Y (NPY). Agrp significantly stimulated light-phase feeding by 24 h post-injection. However, Agrp stimulated dark-phase and deprivation-induced feeding by 4 and 2 h, respectively. Animals receiving NPY and Agrp consumed more than animals receiving either peptide alone, the effect remaining by 24 h.

Evidence of interactions between melanocortin and opioid systems in regulation of feeding

The aim of our experiments was to study the presumed functional relationship between the melanocortin and opioid systems in the regulation of food intake. We determined that a non-selective opioid receptor antagonist, naltrexone, at relatively low doses, decreases food intake induced by i.c.v. agouti-related protein (Agrp). We also observed that peripheral injection of naltrexone at a dose known to produce anorexigenic responses induced c-Fos immunoreactivity in significantly more arcuate nucleus α-MSH neurons than observed in control animals. The results of our study support the notion that the melanocortin and opioid systems interact in the regulation of food intake. Based on these data we speculate that opioid peptides suppress α-MSH-dependent satiety mechanisms; conversely, it is possible that the orexigenic action of Agrp is mediated via opioid dependent circuitry.

Interrelationships between μ opioid and melanocortin receptors in mediating food intake in rats

Abstract The present study examined the interrelationships between feeding responses produced by μ opioid receptor agonists and melanocortin-3 or 4 (MC-3/4) receptor antagonists. Feeding induced by the μ-sensitive opioid peptide, β-endorphin (βEND, 10 μg, icv) was significantly and dose-dependently reduced by pretreatment with the MC-3/4 receptor agonist, melanotan-II (MTII: 0.01–10 nmol, icv). Moreover, the selective μ opioid antagonist, β-funaltrexamine (βFNA: 2–20 μg, icv), significantly and dose-dependently reduced feeding and weight gain elicited by the potent MC-3/4 receptor antagonist, SHU-9119 (0.5 nmol, icv), especially at those intake periods (24–48 h) when SHU-9119 produced maximal ingestive effects. These data extend previous findings demonstrating interactions between opioid and melanocortin receptors in the mediation of food intake.

Direct effects of nutrients, acetylcholine, CCK, and insulin on ghrelin release from the isolated stomachs of rats

Ghrelin is a powerful orexigenic peptide predominantly secreted by the stomach. Blood concentration of ghrelin increases before meals and fall postprandial. Its regulation appears to be influenced by the type of macronutrient ingested, the vagus nerve stimulation and by other post-meal stimulated hormonal factors. However, the direct role of nutrients (amino acids or lipids), neuronal (vagal neurotransmitter acetylcholine) and satiety-inducing factor such as CCK are not known. To study this we applied amino acids, lipids, acetylcholine and CCK via vascular perfusion to the isolated stomachs and found that amino acids significantly reduced ghrelin release from the isolated stomach by approximately approximately 30% vs. the control while lipids (10% intralipid) had no affect. Acetylcholine (1 microM) increased ghrelin release from the stomach by approximately 37% whereas insulin (10nM) decreased it by approximately 30% vs. the control. Interestingly, CCK (100 nM) potently increased ghrelin release by approximately 200% vs. the control. Therefore it appears that ghrelin secretion from the stomach is under direct influence of amino acids, neurotransmitter acetylcholine and hormones such as insulin and CCK.

Effect of reducing hypothalamic ghrelin receptor gene expression on energy balance

Central and peripheral injections of fghrelin potently stimulates food intake via its receptor, GHSR1a expressed in the brain. In this study, we explored the role of GHSR1a in the paraventricular nucleus of the hypothalamus (PVN) by reducing their gene expression using the RNA interference (RNAi). pSUPER plasmids inserted with sh (short hairpin)-GHSR1a were injected into the PVN to reduce its expression. The transfected rats were monitored daily for their food intake and body weight throughout the experimental period lasting 8 days. We found that knockdown of GHSR1a did not affect daily food intake but significantly reduced body weight and blood ghrelin levels. This suggests that the central ghrelin system could selectively regulate body weight without affecting energy intake.

Maternal high fat feeding and gestational dietary restriction: effects on offspring body weight, food intake and hypothalamic gene expression over three generations in mice.

Excessive gestational weight gain and maternal obesity have both been associated with increased incidence of obesity and metabolic disorder in offspring in both humans and animal models. The objectives of this study were to determine (1) whether mild gestational food restriction during the third trimester (GFR) would alter food intake and growth parameters of offspring, (2) whether effects of GFR depended on diet (high fat [HF] vs chow), (3) whether effects of excessive gestational weight gain (WG) would become magnified across generations, and (4) whether diet and GFR would alter hypothalamic gene expression in adult offspring. Three generations of female C57BL/6 mice were fed chow or HF diet, mated at 11 weeks of age and assigned to ad libitum feeding or 25% GFR. Offspring were fed the same diet as their mothers. Results showed (1) maternal gestational WG was positively correlated with offspring WG. (2) HF offspring weighed less (p<0.01) at weaning (WWT) but gained more during the 8 weeks after weaning than chow-fed offspring (p<0.05), resulting in higher final body weights (BW) (p<0.01). (3) HF males from GFR mothers had higher WWT (p<0.05), but subsequent WG and final BW were less (p<0.05) compared to males from ad lib mothers. (4) In the HF group, GFR also resulted in decreased FI (p<0.05) and FE (p<0.07) in offspring, compared to offspring from ad lib mothers. (5) In generation 3, hypothalamic expression of tyrosine hydroxylase was lower in HF males from GFR mothers compared to HF males from ad lib mothers (p<0.05). In conclusion, gender and maternal GFR had independent effects on growth and FI, and hypothalamic gene expression was dependent on both gender and maternal GFR in HF offspring. Even mild food restriction of obese mothers during pregnancy may have beneficial effects in reducing the risk or degree of obesity in offspring.