Reversible suppression of food reward behavior by chronic mu-opioid receptor antagonism in the nucleus accumbens.

Overindulgence in easily available energy-dense palatable foods is thought to be an important factor in the current obesity epidemic but the underlying neural mechanisms are not well understood. Here we demonstrate that mu-opioid receptor signaling in the nucleus accumbens may be important. Protracted suppression of endogenous mu-opioid receptor signaling focused on the nucleus accumbens shell for several days by means of microinjected beta-funaltrexamine (BFNA) diminished both "liking" of sucrose, as indicated by fewer positive hedonic orofacial responses, and the incentive reinforcement value ("wanting") of a food reward, as indicated by lower completion speed and increased time being distracted in the incentive runway. BFNA-treatment also decreased responding to sucrose and corn oil in the brief access lick paradigm, a test measuring a combination of mainly taste-guided "liking" and low-effort "wanting", as well as 4 h intake of sucrose solution. These effects were not due to nonspecific permanent neuronal changes, as they were fully reversible. We conclude that endogenous mu-opioid signaling in the nucleus accumbens is necessary for the full display of palatable food-induced hyperphagia through mechanisms including hedonic, motivational, and reinforcement processes. Development of obesity could be the result of predisposing innate differences in these mechanisms or overstimulation of these mechanisms by external factors.

Food-related gastrointestinal signals activate caudal brainstem neurons expressing both NMDA and AMPA receptors.

Vagal mechano- and chemosensors in the gastrointestinal tract and the portal-hepatic axis signaling the arrival of nutrients are major determinants of the satiation process. Although glutamate and its various receptor subtypes have been shown to transmit gustatory and cardiovascular sensory information at the level of the solitary nucleus (nucleus tractus solitarius; NTS), their involvement in the transmission of gastrointestinal satiety signals is not clear. Gastrointestinal sensors were stimulated by gastric balloon distension or by intraduodenal infusion of either linoleic acid or glucose in chronically catheterized, non-anesthetized rats, leading to activation of second order neurons in the NTS as detected by c-Fos immunohistochemistry. Subsequent (double)-immunohistochemistry for either NMDA or AMPA glutamate receptors was used to determine the percentage of activated neurons expressing a particular receptor subtype. Gastric distension and duodenal nutrient stimuli produced slightly, but significantly different patterns of c-Fos induction in the dorsal vagal complex. Expression of NMDA receptors, as detected by a NR2ab subunit-specific antibody, was abundant throughout the dorsal medulla. The percentage of neurons in the NTS activated by gastric distension (63.9+/-2.9%), linoleic acid (62.8+/-1.4%), and glucose (64.1+/-1.4%), expressing NMDA receptor was similar. Expression of AMPA receptors, as detected by a GLUR2/3 subunit-specific antibody, was equally abundant throughout the dorsal medulla. Again, the percentage of activated neurons expressing GLUR2/3 was similar for the gastric distension (59.8-65.6%) and duodenal linoleic acid (60.6-67.0%) stimuli, and for the various subnuclei of the NTS. Finally, GLUR1-specific immunoreactivity was much less abundant, with only a small percentage of distension-activated (4.4+/-0.4%) and linoleic acid-activated (5.1+/-0.4%) neurons expressing this receptor subunit. The results suggest a widespread, general involvement of both NMDA and AMPA receptors in primary afferent signal transmission at the level of the NTS, with no differential recruitment of the examined receptor subtypes by the different gastrointestinal sensory stimuli. This may indicate a high degree of convergence among sensory signals, or alternatively, the presence of other transmission systems such as peptides referring sensory specificity to second order neurons.

Duodenal nutrient infusions differentially affect sham feeding and Fos expression in rat brain stem.

Duodenal infusions of macronutrients inhibit sham and normal feeding. Neural substrates of this response were studied by infusing glucose, linoleic acid, an amino acid mixture, saline, or water into the duodenum of unanesthetized rats and then measuring sham feeding of 30% sucrose or Fos expression in the dorsal vagal complex. Linoleic acid and amino acids (both 1.5 kcal) and glucose (4.5 kcal) suppressed sham feeding relative to control infusions, and all three macronutrients triggered Fos expression in the nucleus of the solitary tract and area postrema. Although there were significant quantitative differences, the subnuclear distribution pattern of Fos-expressing neurons was not different for the three macronutrients and was largely localized to the medial, dorsomedial, and commissural subnuclei of the nucleus of the solitary tract and the area postrema. Linoleic acid suppressed intake and stimulated Fos expression similarly to glucose infusions of three times the caloric value. Amino acids strongly suppressed sham feeding but triggered relatively little Fos expression. These results indicate that the intake-suppressing potency of duodenal macronutrients is dependent on nutrient type, rather than simply caloric value, and that amino acids, although potent inducers of satiety, affect ingestion by processes different from those subserving lipids and carbohydrates. Furthermore, the similar patterns of neuronal activation after different duodenal infusions may indicate a large degree of convergence at the level of primary and second-order sensory neurons, whereas the distinctly different pattern obtained earlier with gastric distension indicates partially separate neural pathways for satiety signals generated by duodenal nutrients and gastric mechanoreceptors.

The early presence of food-oriented appetitive behavior in developing rats.

The appetitive behavior of 3- to 6-day-old rat pups was studied by testing their ability to direct their ingestive behavior to a restricted food source. We found that, from 3 days of age, pups were able to feed efficiently from such a source. More specifically, pups that were deprived of nutrition but not of maternal care as well as pups that were dehydrated ingested significantly more than nondeprived animals, and did so whether liquid diet was spread over the entire floor surface beneath them or restricted to a fraction of the floor surface. However, pups that had been nutritionally and maternally deprived were not able to direct their feeding. The general locomotor activation of pups in this latter group appeared to interfere with their ability to direct their behavior to the restricted source. These results indicate that from early ages, developing rats possess the appetitive competence to guide their behavior and suggest that previous findings of poorly directed behavior were a confound of the behavioral activation shown by pups tested in a state of maternal deprivation.

Effects of hydrational state on ingestion in infant rats: is dehydration the only ingestive stimulus?

The physiological stimulus for deprivation-enhanced ingestion was studied in developing rats. During an overnight deprivation period, continuous gastric infusions of isotonic saline or milk were made to 6- and 15-day-old rat pups in order to preferentially maintain hydrational or hydrational and nutritional status, respectively. Pups' ingestion was then studied in oral-infusion tests. In 6-day-old pups that received either milk or saline infusions, ingestion was depressed relative to intake in pups that were simply deprived. But in 15-day-old pups, only milk infusions reduced intake. These findings suggest that the increased ingestion stimulated by deprivation in pups less than a week of age results primarily from dehydration, and thus that nutrient-related feeding does not emerge until later in development.

Ingestive behavior in preweanling rats: emergence of postgastric controls.

Previous studies have indicated that the termination of intake in very young rat pups is controlled almost exclusively by the level of gastric fill; nutritive cues from diet have no effect on intake. In the present series of experiments, we investigated the ontogeny of postgastric nutritive controls of intake in rat pups ingesting independent of their dam. In 6-day-old pups, the level of gastric fill required to terminate ingestion was not affected by the presence or absence of post-gastric nutritive cues, but by 15 days of age a greater level of gastric fill was required to stop ingestion when postgastric cues were eliminated by a closed pyloric noose. This emergent post-gastric contribution to the inhibition of intake in 15-day-old pups does not depend on preexisting gastric fill signals, as sham-feeding pups (open gastric fistula) with nutrients in their intestines ingested less than pups without intestinal nutrients. These results provide evidence that postgastric controls of ingestive behavior mature during the postnatal period and just shortly before they are required at weaning.

Ontogeny of glucose inhibition of independent ingestion in preweanling rats.

Rat pups that have been maternally and nutritionally deprived will vigorously ingest diet infused directly into the mouth. The development of nutritive controls in this form of ingestion was examined by administering nutritive and non-nutritive gastric preloads to 6- and 15-day-old pups. In 6 day-old pups, nutritive gastric preloads (0.6 M glucose in distilled H2O or saline) and vehicle preloads were followed by similar intakes; only the change in hydrational state caused by distilled H2O loads appeared to affect intake. By 15 days of age, intake following nutritive preloads was less than intake following non-nutritive preloads. Also, at 15 days, stomach volume at the termination of intake was less following nutritive preloads. In a separate experiment with 6-day-old pups, gastric preloads of an alternative energy source, the ketone beta-hydroxybutyrate, also failed to inhibit intake when given at a dose that did not cause excessive gastric distension. These results indicate that a nutritive control of intake termination in rats is not present at 6 days of age but develops by 15 days of age.

Control of ingestion in 6-day-old rat pups: termination of intake by gastric fill alone?

Six-day-old rat pups, tested away from the dam, vigorously ingested orally infused milk for up to 90 min when the ingested milk was allowed to drain through a gastric fistula. Total intake in pups with unplugged fistulas was more than double that consumed by littermates with plugged fistulas. This persistent feeding could not be due to the absence of postgastric cues because blocking movement of the milk into the intestine with a closed pyloric noose resulted in less intake than when the noose was open. Furthermore, a 5% body weight gastric preload of milk or nonnutritive saline suppressed intake in pups with either open or closed pyloric nooses by an amount equal to the preload. Regardless of noose or preload condition, ingestion stopped when stomach content reached approximately 6.5% of body weight. Thus level of gastric fill appeared critical for intake control. When gastric fistulas were unplugged after pups had ingested milk until they stopped, vigorous ingestion followed, with total intakes of up to 20% body weight. Intake termination in young rat pups thus appears to be controlled exclusively by gastric fill level; pregastric signals alone appear insufficient, and postgastric signals appear unnecessary for termination of intake.

Use of hypothermia for general anesthesia in preweanling rodents.

Documentation and rationale are provided for the use of deep hypothermia alone for anesthesia during surgical manipulations in young preweanling rodents and moderate hypothermia as an adjunct to other anesthesia in older preweanling animals. Techniques and applications for the procedure are also described.

Body hydration and haemolymph osmolality affect feeding and its neural correlate in the terrestrial gastropod, Limax maximus.

When terrestrial slugs (Limax maximus) are dehydrated to 65-70% of their initial body weight (IBW) their feeding responsiveness is greatly decreased. There is a 90% decrease in feeding responsiveness when slugs are injected with hyperosmotic mannitol solution that raises the haemolymph osmolality to that of slugs dehydrated to 65-70% IBW (i.e. 200 mosmol kg-1 H2O). The duration of the Feeding Motor Programme (FMP) that can be recorded from an isolated CNS-lip preparation is reduced by increasing the osmolality of the saline bathing the preparation. The osmolality of the saline that can modify the FMP corresponds to that of the haemolymph of a slug dehydrated to 65-70% IBW. The pattern of the motor programme is not affected. A gradual increase in saline osmolality which temporally mimics the progressive increase in haemolymph osmolality of a dehydrating slug also causes a decrease in the duration of the FMP. The neural network underlying the FMP appears to adapt to hyperosmotic saline since the duration of FMP bouts gradually returns to normal levels after long-term exposure (6-8 h).