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These studies investigated feeding responses to indirect activation of parabrachial cannabinoid CBl receptors. Arachidonoyl serotonin (AA5HT), an inhibitor of the endocannabinoid degradative enzyme, fatty acid amide hydrolase (FAAH), was infused into the parabrachial nucleus of male Sprague-Dawley rats, and intakes of high-fat/sucrose pellets and standard rodent chow were subsequently evaluated under various feeding schedules. FAAH blockade stimulated the intake of high-fat/sucrose pellets that were presented daily for 4 h during the light period, with compensatory decreases in the consumption of standard chow during the ensuing 20 h. These diet-selective changes were repeated on the next day, indicating a shift in feeding toward the more palatable diet that lasted for 48 h after a single infusion. The cannabinoid CBI receptor antagonist, AM251, blocked the orexigenic actions of AA5HT, implicating CBI receptors in mediating the feeding responses to FAAH inactivation. When the feeding schedule was reversed, AA5HT produced nominal increases in the consumption of standard chow for the 4-h access period, but substantial increases in the intake of high-fat/sucrose during the following 20-h interval. When presented with only high-fat/sucrose diet for 24 h, AA5HT increased 24-h food intake. In contrast, when given 24-h access only to standard chow, AA5HT failed to affect intake. Therefore, indirectly activating parabrachial CB 1 receptors by blocking the degradation of native ligands selectively stimulates the intake of palatable food, with differential actions on total energy intake depending upon the feeding schedule. Our results support a role for parabrachial cannabinoid mechanisms in providing physiological regulation to neural substrates modulating feeding, energy balance, and behavioral responses for natural reward. parabrachial nucleus; endocannahinoids; feeding; reward

Wilson, John D., Danielle M. Nicklous, Vincent J. Aloyo, and Kenny J. Simansky. An orexigenic role for [micro]-opioid receptors in the lateral parabrachial nucleus. Am J Physiol Regul Integr Comp Physiol 285: R1055-R1065, 2003; 10.1152/ajpregu.00108.2003. The pontine parabrachial nucleus (PBN) has been implicated in regulating ingestion and contains opioids that promote feeding elsewhere in the brain. We tested the actions of the selective [micro]-opioid receptor ([micro]-OR) agonist [D-[Ala.sup.2],N-Me-[Phe.sup.4],[Gly.sup.5]_ol]enkephalin (DAMGO) in the PBN on feeding in male rats with free access to food. Infusing DAMGO (0.5-4.0 nmol/0.5 [micro]l) into the lateral parabrachial region (LPBN) increased food intake. The hyperphagic effect was anatomically specific to infusions within the LPBN, dose and time related, and selective for ingestion of chow compared with (nonnutritive) kaolin. The nonselective opioid antagonist naloxone (0.1-10.0 nmol intra-PBN) antagonized DAMGO-induced feeding, with complete blockade by 1.0 nmol and no effect on baseline. The highly selective [micro]-opioid antagonist D-Phe-Cys-Trp-Arg-Thr-Pen-Thr-[NH.sub.2] (CTAP; 1.0 nmol) also prevented this action of DAMGO, but the [kappa]-antagonist nor-binaltorphimine did not. Naloxone and CTAP (10.0 nmol) decreased intake during scheduled feeding. Thus stimulating [micro]-ORs in the LPBN increases feeding, whereas antagonizing these sites inhibits feeding. Together, our results implicate [micro]-ORs in the LPBN in the normal regulation of food intake. feeding; hyperphagia; DAMGO; naloxone; CTAP

Nicklous, Daniele M. and Kenny J. Simansky. Neuropeptide FF exerts pro- and anti-opioid actions in the parabrachial nucleus to modulate food intake. Am J Physiol Regul Integr Comp Physiol 285: R1046-R1054, 2003; 10.1152/ajpregu.00107.2003. --Neurons that synthesize the morphine modulatory peptide neuropeptide FF (NPFF; Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2) densely innervate the parabrachial nucleus (PBN), an area implicated in regulating food intake. We analyzed opioid-related actions of NPFF in feeding in adult male Sprague-Dawley rats. Unilateral infusion of 2 nmol/0.5 [micro]1 of the [micro]-opioid receptor agonist [D-[Ala. sup.2],N-Me-[Phe.sup.4], [glycinol.sup.5]] enkephalin (DAMGO) into the lateral PBN increased 4-h food intake from 0.7 [+ or -] 0.1 to 3.3 [+ or -] 0.3 g. NPFF (1.25-5.0 nmol) prevented this hyperphagic [micro]-opioidergic action. In rats fed after 4-h deprivation (baseline = 12.3 [+ or -]0.3 g/2 h), 5 nmol of NPFF did not alter and larger doses (10 and 20 nmol) actually increased food intake (+36, 54%). Twenty nanomoles also elevated intake of freely feeding rats (from 0.7 [+ or -] 0.1 to 5.1 [+ or -] 1.0 g/4 h). The opioid receptor blocker naloxone (10 nmol) antagonized this increase. These data reveal both pro- and anti-opioid actions of NPFF in the PBN to modulate feeding. The mechanisms for the opposite actions of low and high concentrations of this neuropeptide in parabrachial regulation of food intake remain to be determined. feeding; hyperphagia; rats; FMRFamide; [micro]-opioid receptors

A study was conducted to compare the influence of total abdominal vagotomy on the decrease in food intake caused by serotonergic (5-HTergic) and cholecystokinergic (CCKergic) stimulation in rats. The results revealed that 5-carboxamidotryptamine (5-CT), produces anorexia through a vagally independent function, but stimulates drinking by a pathway which does not include vagal function. The theory that vagotomy disintegrates the neural functions for the anorectic working of peripheral 5-HTergic and CCKergic stimulation was proved.

The endocannabinoid system is emerging as an integral component in central and peripheral regulation of feeding and energy balance. Our investigation analyzed behavioral roles for cannabinoid mechanisms of the pontine parabrachial nucleus (PBN) in modulating intake of presumably palatable foods containing fat and/or sugar. The PBN serves to gate neurotransmission associated with, but not limited to, the gustatory properties of food. Immunofluorescence and in vitro [35S]GTPγS autoradiography of rat tissue sections containing the PBN revealed the presence of cannabinoid receptors and their functional capability to couple to their G-proteins after incubation with the endocannabinoid 2-arachidonoyl glycerol (2-AG). The selective cannabinoid 1 receptor (CB1R) antagonist AM251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] prevented the response, demonstrating CB1R mediation of 2-AG-induced coupling. Microinfusions of 2-AG into the PBN in behaving rats robustly stimulated feeding of pellets high in content of fat and sucrose (HFS), pure sucrose, and pure fat (Crisco), during the first 30 min after infusion. In contrast, 2-AG failed to increase consumption of standard chow, even when the feeding regimen was manipulated to match baseline intakes of HFS. Orexigenic responses to 2-AG were attenuated by AM251, again indicating CB1R mediation of 2-AG actions. Furthermore, responses were regionally specific, because 2-AG failed to alter intake when infused into sites ∼500 μm caudal to infusions that successfully stimulated feeding. Our data suggest that hedonically positive sensory properties of food enable endocannabinoids at PBN CB1Rs to initiate increases in eating, and, more generally, these pathways may serve a larger role in brain functions controlling behavioral responses for natural reward. [ABSTRACT FROM AUTHOR]

Serotonergic 5-HT2C and 5-HT1B receptors mediate inhibitory controls of eating. Questions have arisen about potential behavioral and neurological toxicity of drugs that stimulate the 2C site. We evaluated eating and other motor responses in male Dutch-belted rabbits after administration of m-chlorophenylpiperazine (mCPP). Studies conducted in vitro and in vivo assessed the pharmacological specificity of the ingestive actions of this agent. mCPP (0.15–10 μmol/kg sc) reduced consumption of chow and 20% sucrose solution with equal potencies (ED500.6 μmol/kg). In radioligand binding to rabbit cortex, mCPP displayed 15-fold higher affinity for 5-HT2C than for 5-HT1B receptors. The serotonin antagonist mesulergine (7000-fold selective for 5-HT2C) reversed the hypophagic action of mCPP, but the 5-HT1B/1D antagonist GR127,935 did not. GR127,935 (0.5 μmol/kg) did prevent hypophagia produced by the highly selective 5-HT1B/1D agonist GR46,611. Observational methods demonstrated that mCPP decreased the frequency of eating chow but increased other motor activities. When rabbits consumed sucrose, videoanalysis revealed that mCPP reduced total time licking and the duration of individual bouts, but not bout frequency or the actual rate of consumption. mCPP increased locomotor and other activities, and greatly increased vacuous oromotor stereotypies and tongue protrusions. Nonetheless, rabbits licked accurately at the spout for sucrose. When sucrose was infused intraorally through a cheek catheter, mCPP actually increased the peak amplitude and overall magnitude of jaw movements. We conclude that mCPP stimulates 5-HT2C receptors to reduce food intake in rabbits. This hypophagia involves disruption of appetitive components of eating and is accompanied by adverse motor actions. This profile raises questions about the use of the 5-HT2C receptor as a target for novel therapeutic agents for obesity. [Copyright &y& Elsevier]

Systemic administration of the serotonin (5-HT) releaser/reuptake inhibitor, d-fenfluramine decreases consumption of food in mammals. This hypophagic action involves loci at several levels of the neuraxis. Indirect evidence implicates the parabrachial nucleus (PBN) of the pons as one of these regions. Consistent with this hypothesis, unilateral infusion of d-fenfluramine (200, 280, and 400 nmol/0.5 μl) directly into the lateral PBN (LPBN) of male rats reduced food intake by 33%, 56%, and 66% from baseline (7.3±0.7 g) during a 30-min test with chow. Infusions lateral, medial, and dorsal to the PBN were ineffective. Stimulating 5-HT1B receptors in the PBN also reduces feeding. Administration of the selective 5-HT1B agonist CP-93,129 (3-(1,2,5,6-tetrahydropyrid-4-yl)pyrrolo[3,2-b]pyrid-5-one) (0, 0.625, 2.5, and 10 nmol/0.5 μl) into the PBN reduced food intake by 25–79%. The selective 5-HT1B antagonist SB-216641 (N-[3-[3-(dimethylamino(ethoxy]-4-methoxyphenyl]-2′-methyl-4′-(5-methyl-1,2,4-oxadiazol-3-yl)-[1,1′-biphenyl]-4-carboxamide) (2.5 nmol) completely blocked the hypophagic action of the approximate ED50 doses of CP-93,129 (2.5 nmol) and d-fenfluramine (280 nmol). These data strongly suggest that directly or indirectly activating 5-HT1B receptors in the LPBN inhibits feeding and implicates this pontine region in the serotonergic regulation of eating and satiation. [Copyright &y& Elsevier]

Abstract: The parabrachial nucleus (PBN) is an area of the brain stem that controls eating and contains endogenous opioids and their receptors. Previously, we demonstrated that acute activation of μ opioid receptors (MOPR) in the lateral PBN increased food consumption. MOPRs have been divided operationally into μ1 and μ2 receptor subtypes on the basis of the ability of naloxonazine (Nlxz) to block the former but not the latter. We used autoradiography to measure whether Nlxz blocks stimulation by the μ1/μ2 agonist DAMGO (D-Ala2, N-Me-Phe4, Gly5-ol-enkephalin) of the incorporation of [35S]-guanosine 5′(γ-thio)triphosphate ([35S]-GTPγS) into sections of the PBN. In vitro, Nlxz dose dependently inhibited receptor coupling in all areas of the PBN. The 1 μM concentration of Nlxz reduced stimulation by 93.1±5% in the lateral inferior PBN (LPBNi) and by 90.5±4% in the medial parabrachial subregion (MPBN). Administration of Nlxz directly into the LPBNi decreased both food intake and agonist stimulated coupling, ex vivo, for the 24-h period after infusion. Infusion of Nlxz into the intended area reduced food intake by 42.3% below baseline values. Nlxz infusion prevented DAMGO stimulation of G-protein coupling in LPBNi and markedly reduced this stimulation in the MPBN. The incomplete inhibition of DAMGO-stimulated coupling in the MPBN is most likely due to the limited diffusion of Nlxz from the site of infusion (LPBNi) into this brain region. In conclusion, this study demonstrates that the μ1 opioid receptor subtype is present in the parabrachial nucleus of the pons and that these receptors serve to modulate feeding in rats. [Copyright &y& Elsevier]

The parabrachial nucleus (PBN) is an area of the brain stem that controls eating and contains endogenous opioids and their receptors. Previously, we demonstrated that acute activation of mu opioid receptors (MOPR) in the lateral PBN increased food consumption. MOPRs have been divided operationally into mu(1) and mu(2) receptor subtypes on the basis of the ability of naloxonazine (Nlxz) to block the former but not the latter. We used autoradiography to measure whether Nlxz blocks stimulation by the mu(1)/mu(2) agonist DAMGO (D-Ala2, N-Me-Phe4, Gly5-ol-enkephalin) of the incorporation of [(35)S]-guanosine 5'(gamma-thio)triphosphate ([(35)S]-GTPgammaS) into sections of the PBN. In vitro, Nlxz dose dependently inhibited receptor coupling in all areas of the PBN. The 1 muM concentration of Nlxz reduced stimulation by 93.1+/-5% in the lateral inferior PBN (LPBNi) and by 90.5+/-4% in the medial parabrachial subregion (MPBN). Administration of Nlxz directly into the LPBNi decreased both food intake and agonist stimulated coupling, ex vivo, for the 24-h period after infusion. Infusion of Nlxz into the intended area reduced food intake by 42.3% below baseline values. Nlxz infusion prevented DAMGO stimulation of G-protein coupling in LPBNi and markedly reduced this stimulation in the MPBN. The incomplete inhibition of DAMGO-stimulated coupling in the MPBN is most likely due to the limited diffusion of Nlxz from the site of infusion (LPBNi) into this brain region. In conclusion, this study demonstrates that the mu(1) opioid receptor subtype is present in the parabrachial nucleus of the pons and that these receptors serve to modulate feeding in rats.

The endocannabinoid system is emerging as an integral component in central and peripheral regulation of feeding and energy balance. Our investigation analyzed behavioral roles for cannabinoid mechanisms of the pontine parabrachial nucleus (PBN) in modulating intake of presumably palatable foods containing fat and/or sugar. The PBN serves to gate neurotransmission associated with, but not limited to, the gustatory properties of food. Immunofluorescence and in vitro [(35)S]GTPgammaS autoradiography of rat tissue sections containing the PBN revealed the presence of cannabinoid receptors and their functional capability to couple to their G-proteins after incubation with the endocannabinoid 2-arachidonoyl glycerol (2-AG). The selective cannabinoid 1 receptor (CB(1)R) antagonist AM251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] prevented the response, demonstrating CB(1)R mediation of 2-AG-induced coupling. Microinfusions of 2-AG into the PBN in behaving rats robustly stimulated feeding of pellets high in content of fat and sucrose (HFS), pure sucrose, and pure fat (Crisco), during the first 30 min after infusion. In contrast, 2-AG failed to increase consumption of standard chow, even when the feeding regimen was manipulated to match baseline intakes of HFS. Orexigenic responses to 2-AG were attenuated by AM251, again indicating CB(1)R mediation of 2-AG actions. Furthermore, responses were regionally specific, because 2-AG failed to alter intake when infused into sites approximately 500 mum caudal to infusions that successfully stimulated feeding. Our data suggest that hedonically positive sensory properties of food enable endocannabinoids at PBN CB(1)Rs to initiate increases in eating, and, more generally, these pathways may serve a larger role in brain functions controlling behavioral responses for natural reward.

Neurons that synthesize the morphine modulatory peptide neuropeptide FF (NPFF; Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2) densely innervate the parabrachial nucleus (PBN), an area implicated in regulating food intake. We analyzed opioid-related actions of NPFF in feeding in adult male Sprague-Dawley rats. Unilateral infusion of 2 nmol/0.5 microl of the mu-opioid receptor agonist [d-Ala2,NMe-Phe4,glycinol5]enkephalin (DAMGO) into the lateral PBN increased 4-h food intake from 0.7 +/- 0.1 to 3.3 +/- 0.3 g. NPFF (1.25-5.0 nmol) prevented this hyperphagic mu-opioidergic action. In rats fed after 4-h deprivation (baseline = 12.3 +/- 0.3 g/2 h), 5 nmol of NPFF did not alter and larger doses (10 and 20 nmol) actually increased food intake (+36, 54%). Twenty nanomoles also elevated intake of freely feeding rats (from 0.7 +/- 0.1 to 5.1 +/- 1.0 g/4 h). The opioid receptor blocker naloxone (10 nmol) antagonized this increase. These data reveal both pro- and anti-opioid actions of NPFF in the PBN to modulate feeding. The mechanisms for the opposite actions of low and high concentrations of this neuropeptide in parabrachial regulation of food intake remain to be determined.

The pontine parabrachial nucleus (PBN) has been implicated in regulating ingestion and contains opioids that promote feeding elsewhere in the brain. We tested the actions of the selective mu-opioid receptor (mu-OR) agonist [d-Ala2,N-Me-Phe4,Gly5-ol]enkephalin (DAMGO) in the PBN on feeding in male rats with free access to food. Infusing DAMGO (0.5-4.0 nmol/0.5 microl) into the lateral parabrachial region (LPBN) increased food intake. The hyperphagic effect was anatomically specific to infusions within the LPBN, dose and time related, and selective for ingestion of chow compared with (nonnutritive) kaolin. The nonselective opioid antagonist naloxone (0.1-10.0 nmol intra-PBN) antagonized DAMGO-induced feeding, with complete blockade by 1.0 nmol and no effect on baseline. The highly selective mu-opioid antagonist d-Phe-Cys-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP; 1.0 nmol) also prevented this action of DAMGO, but the kappa-antagonist nor-binaltorphimine did not. Naloxone and CTAP (10.0 nmol) decreased intake during scheduled feeding. Thus stimulating mu-ORs in the LPBN increases feeding, whereas antagonizing these sites inhibits feeding. Together, our results implicate mu-ORs in the LPBN in the normal regulation of food intake.