Hydrolytic instability of the important orexin 1 receptor antagonist SB-334867: Possible confounding effects on in vivo and in vitro studies

SB-334867 has been an important ligand for the study of the orexin 1 (OX1) receptor due to its high OX1/OX2 selectivity and bioavailability. This ligand however, contains a 2-methylbenzoxazole ring system which is known to undergo hydrolysis, particularly under acidic or basic conditions. The possibility that SB-334867 would be susceptible to significant hydrolysis was evaluated in various formulations and in the solid state. SB-334867 was found to be unstable under conditions commonly employed to prepare stock solutions for in vitro and in vivo studies. In addition, and most alarmingly, the hydrochloride salt of SB-334867 was found to quantitatively decompose to an OX1-inactive product even in the solid state. These findings combine to suggest that studies using SB-334867 (and any other 2-methylbenzoxazole-containing compound) should be performed with great care to avoid the confounding effects of the rapid hydrolytic decomposition of this susceptible structure.     

Anorectic, thermogenic and anti-obesity activity of a selective orexin-1 receptor antagonist in ob/ob mice

A single dose of the orexin-1 (OX1) receptor antagonist 1-(2-methylbenzoxazol-6-yl)-3-[1,5] naphthyridin-4-yl urea hydrochloride (SB-334867-A) reduces orexin-A-induced feeding and natural feeding in Sprague Dawley rats. In this study, the anti-obesity effects of SB-334867-A were determined in genetically obese (ob/ob) mice dosed with SB-334867-A (30 mg/kg, i.p.) once daily for 7 days, and then twice daily for a further 7 days. SB-334867-A reduced cumulative food intake and body weight gain over 14 days. Total fat mass gain, determined by Dual Emission X-ray Absorptiometry, was reduced, while gain in fat-free mass was unchanged. Fasting (5 h) blood glucose was also reduced at the end of the study, with a trend to reduced plasma insulin. Interscapular brown adipose tissue (BAT) weight was reduced, the tissue was noticeably darker in colour and quantitative PCR (TaqMan) analysis of this tissue showed a trend to an increase in uncoupling protein-1 mRNA expression, suggesting that SB-334867-A might stimulate thermogenesis. This was confirmed in a separate study in which a single dose of SB-334867-A (30 mg/kg, i.p.) increased metabolic rate over 4 h in ob/ob mice. OX1 receptor mRNA was detected in BAT, and its expression was increased by 58% by treatment with SB-334867-A. This is the first demonstration that OX1 receptor antagonists have potential as both anti-obesity and anti-diabetic agents.     

The selective orexin 1 receptor antagonist SB-334867-A impairs acquisition and consolidation but not retrieval of spatial memory in Morris water maze

The novel neuropeptides orexin-A and orexin-B derive from a common 130-amino acid precursor molecule (prepro-orexin), are mainly localized to neurons within and around the lateral hypothalamus, and exhibit high affinity to the closely related G-Protein-coupled receptors orexin 1 and 2 receptor (OX1R, OX2R). Orexinergic neurons send their axons to the hippocampal formation (CA1, CA2 and dentate gyrus), which expresses OX1Rs. Recent studies have shown that central administration of orexin-A and orexin-B have effects on learning and memory but literature concerning the role of orexinergic system in cognition remains controversial. More recently, antagonists have been described. The most potent and selective is SB-334867-A, which has an affinity of 40 nM at OX1R which is at least 50-fold selective over OX2R. It is likely that the intracerebroventricular (i.c.v.) administration may block OX1Rs in many brain regions. Previously we have shown that intra-CA1 injection of SB-334867-A impairs acquisition, consolidation and retrieval of spatial memory in MWM task. In the present study, the effect of pre-training, post-training and pre-probe of trial intra-DG (dentate gyrus) administration of SB-334867-A (1.5, 3, 6 microg/0.5 microl) on acquisition, consolidation and retrieval in a single-day testing version of MWM (Morris water maze) task was examined. Our results show impaired acquisition and consolidation of MWM task for SB-334867-A as compared with the control group. However, SB-334867-A had no effect on retrieval in spatial memory. Also, this antagonist had no effect on escape latency of a non-spatial visual discrimination task. Therefore, it seems that endogenous orexin-A and orexin-B, through DG OX1Rs, play an important role in spatial learning and memory in the rat.     

Stimulatory effect of endogenous orexin A on gastric emptying and acid secretion independent of gastrin

Orexin A (OXA) increases food intake and inhibits fasting small bowel motility in rats. The aim of this study was to examine the effect of exogenous OXA and endogenous OXA on gastric emptying, acid secretion, glucose metabolism and distribution of orexin immunoreactivity in the stomach. Rats equipped with a gastric fistula were subjected to intravenous (IV) infusion of OXA or the selective orexin-1 receptor (OX1R) antagonist SB-334867-A during saline or pentagastrin infusion. Gastric emptying was studied with a liquid non-nutrient or nutrient, using 51Cr as radioactive marker. Gastric retention was measured after a 20-min infusion of OXA or SB-334867-A. Plasma concentrations of OXA, insulin, glucagon, glucose and gastrin were studied. Immunohistochemistry against OXA, OX1R and gastrin in gastric tissue was performed. OXA alone had no effect on either acid secretion or gastric emptying. SB-334867-A inhibited both basal and pentagastrin-induced gastric acid secretion and increased gastric retention of the liquid nutrient, but not PEG 4000. Plasma gastrin levels were unchanged by IV OXA or SB-334867-A. Plasma OXA levels decreased after intake of the nutrient meal and infusion of the OX1R antagonist. Only weak effects were seen on plasma glucose and insulin by OXA. Immunoreactivity to OXA and OX1R were found in the mucosa, myenteric cells bodies and varicose nerve fibers in ganglia and circular muscle of the stomach. In conclusion, endogenous OXA influences gastric emptying of a nutrient liquid and gastric acid secretion independent of gastrin. This indicates a role for endogenous OXA, not only in metabolic homeostasis, but also in the pre-absorptive processing of nutrients in the gut.     

Physiological regulation and NO-dependent inhibition of migrating myoelectric complex in the rat small bowel by OXA

Orexin A (OXA)-positive neurons are found in the lateral hypothalamic area and the enteric nervous system. The aim of this study was to investigate the mechanism of OXA action on small bowel motility. Electrodes were implanted in the serosa of the rat small intestine for recordings of myoelectric activity during infusion of saline or OXA in naive rats, vagotomized rats, rats pretreated with guanethidine (3 mg/kg) or N(omega)-nitro-L-arginine (L-NNA; 1 mg/kg). Naive rats were given a bolus of the orexin receptor-1 (OX1R) antagonist (SB-334867-A; 10 mg/kg), and the effect of both OXA and SB-334867-A on fasting motility was studied. Double-label immunocytochemistry with primary antibodies against OXA, neuronal nitric oxide synthase (nNOS), and OX1R was performed. OXA induced a dose-dependent prolongation of the cycle length of the migrating myoelectric complex (MMC) and, in the higher doses, replaced the activity fronts with an irregular spiking pattern. Vagotomy or pretreatment with guanethidine failed to prevent the response to OXA. The OXA-induced effect on the MMC cycle length was completely inhibited by pretreatment with L-NNA (P < 0.05), as did SB-334867-A. The OX1R antagonist shortened the MMC cycle length from 14.1 (12.0-23.5) to 11.0 (9.5-14.7) min (P < 0.05) during control and treatment periods, respectively. Colocalization of OXA and nNOS was observed in myenteric neurons of the duodenum and nerve fibers in the circular muscle. Our results indicate that OXA inhibition of the MMC involves the OX1R and that activation of a L-arginine/NO pathway possibly originating from OX1R/nNOS-containing neurons in the myenteric plexus may mediate this effect. Endogenous OXA may have a physiological role in regulating the MMC.     

Both orexin receptors are expressed in rat ovaries and fluctuate with the estrous cycle: effects of orexin receptor antagonists on gonadotropins and ovulation

Orexins are peptides controlling feeding, sleep, and neuroendocrine functions. They are synthesized by the hypothalamus with projections throughout the brain. Orexins and their orexin 1 (OX(1)) and orexin 2 receptors (OX(2)) are present outside the central nervous system. Here the expression of preproorexin (PPO), OX(1), and OX(2) was studied in rat ovaries. PPO, OX(1), and OX(2) were determined by quantitative real-time RT-PCR in ovaries of cycling Sprague-Dawley rats on all days of the cycle. Serum hormones and food consumption were determined. Ovarian OX(1) and OX(2) expression was then studied after ovulation blockade with Cetrorelix or Nembutal. Finally, proestrous rats were treated at 1400 and 1900 with a selective OX(1) antagonist (SB-334867-A) and/or a selective OX(2) antagonist (JNJ-10397049), and hormone levels, ovulation, and ovarian histology were studied. Both receptors' expression increased in the ovary between 1700 and 2300 of proestrus exclusively, in coincidence with hormone peaks, but not with the dark-light cycle or food intake. PPO was not detected. Cetrorelix or Nembutal prevented the increases of OX(1) and OX(2) while blunting gonadotropin peaks. SB-334867-A and JNJ-10397049, alone or combined, decreased serum gonadotropins and reduced ova number the following morning; ovaries showed a bloody (hyperemic and/or hemorrhagic) reaction with more preovulatory follicles and less corpora lutea. Here we demonstrate for the first time an increased ovarian expression of both OX(1) and OX(2), only during proestrous afternoon, and its hormone dependence but not dependence on the dark-light cycle. Two new receptor antagonists reduced proestrous gonadotropins and/or ova number while producing ovarian structural changes.     

Orexin-A受体介导的生长抑素激动剂ODT8-SST对大鼠摄食和饮水的影响

目的：探讨orexin-A（OXA）受体介导的生长抑素激动剂ODT8-SST 对大鼠摄食和饮水的调节作用相关作用机制。方法：在光 照周期内,大鼠40 只随机分8 组,侧脑室（icv）分别注射不同剂量ODT8-SST 或生理盐水（NS）;大鼠56 只随机分8 组分别侧脑 室注射不同剂量OXA 受体（OX1R）拮抗剂SB-334867 或NS;2小时后测量大鼠摄食量和饮水量。结果：与NS组相比,实验组大 鼠侧脑室注射ODT8-SST（1 ug/rat）,2 小时后摄食量和饮水量均显著增加（P<0.05）。大鼠侧脑室注射SB-334867（16 ug/rat）完全 抑制了由侧脑室注射ODT8-SST 后引起的摄食量和饮水量的增加;与此相反,大鼠给予SST2 拮抗剂S-406-028 预处理之后,可阻 止侧脑室注射ODT8-SST 引发的促进食欲作用,但不会影响侧脑室注射OXA（10.7 ug/rat）诱导的摄食量和饮水量的增加。结论： 侧脑室注射ODT8-SST 可促进摄食和饮水,该过程可能由OX1R所介导;orexin-A 促进摄食作用不依赖大脑SST2 通路的激活。     

The orexin-1 receptor antagonist SB-334867 attenuates anxiety in rats exposed to cat odor but not the elevated plus maze: An investigation of Trial 1 and Trial 2 effects

The orexins are hypothalamic neuropeptides most well known for their roles in regulating feeding and sleeping behaviors. Recent findings suggest that orexin-A may also modulate anxiety, although how and when the orexin system is involved remains unclear. To address this, we investigated the dose-dependent effects of the orexin-1 receptor antagonist SB-334867 in two rodent models of anxiety: the cat odor avoidance model and the elevated plus maze. In both models we tested the effects of SB-334867 when anxiety is novel (Trial 1) and familiar (Trial 2). In the first experiment, Wistar rats were treated with vehicle or SB-334867 (5, 10 or 20 mg/kg, i.p.) prior to their first or second exposure to cat odor. During Trial 1, rats treated with 10 mg/kg of SB-334867 approached the cat odor stimulus more than vehicle-treated rats. During Trial 2 the effects were more marked, with 10 mg/kg of SB-334867 increasing approach times, increasing the number of times rats exited the hide box to engage in exploratory behavior, and decreasing overall hide times. In addition, the 20 mg/kg dose decreased general activity during Trial 2. In the second experiment, the effects of SB-334867 (10 and 20 mg/kg) were tested in the elevated plus maze. There were no significant differences produced by drug treatment during either Trial 1 or Trial 2. Results suggest that SB-334867 decreases anxiety induced by some, but not all, stressors.     

Effect of a selective OX1R antagonist on food intake and body weight in two strains of rats that differ in susceptibility to dietary-induced obesity

An orexin-1 receptor antagonist decreases food intake whereas orexin-A selectively induces hyperphagia to a high-fat diet. In the present study, we evaluated the effect of an orexin antagonist in two strains of rats that differ in their sensitivity to becoming obese while eating a high-fat diet. Male Osborne-Mendel (OM) and S5B/Pl (S5B) rats were treated acutely with an orexin-1 receptor antagonist (SB-334867), after adaptation to either a high-fat (56% fat energy) diet or a low-fat (10% fat energy) diet that were equicaloric for protein (24% energy). Ad libitum fed rats were injected intraperitoneally with SB-334867 at doses of 3, 10 or 30 mg/kg, or vehicle at the beginning of the dark cycle, and food intake and body weight were measured. Hypothalamic prepro-orexin and orexin-1 receptor mRNA expression were analyzed in OM and S5B rats fed at a high-fat or low-fat diet for two weeks. SB-334867 significantly decreased food intake in both strains of rats eating the high-fat diet but only in the OM rats eating the low fat diet. The effect was greatest at 12 and 24 h. Body weight was also reduced in OM rats 1d after injection of SB-334867 but not in the S5B rats. Prepro-orexin and orexin-1 receptor expression levels did not differ between strains or diets. These experiments demonstrate that an orexin antagonist (SB-334867) reduces food intake and has a greater effect in a rat strain that is susceptible to dietary-induced obesity, than in a resistant strain.     

Orexin receptor type-1 antagonist SB-334867 inhibits the development of morphine analgesic tolerance in rats

Herein the effect of orexin receptor type-1 antagonist SB-334867 on the development of tolerance to analgesic effects of morphine was studied in rats. To incite tolerance, morphine sulfate was injected intraperitoneally (i.p., 10mg/kg) once a day for 7 days. The tail flick test was used to evaluate antinociceptive effects of the morphine. A selective OxR1 receptor antagonist, SB-334867, was microinjected (i.c.v.) into the right cerebral ventricle (10 μg/10 μl) immediately before each morphine injection. Repeated morphine application resulted in tolerance to morphine analgesic effects as a decreasing trend during 7 days. Also, repeated administration of SB-334867 (i.c.v.) alone was without significant effect on the nociception as compared to control. Microinjection of SB-334867 prior to each morphine injection inhibited the development of tolerance, so that the analgesic effects of morphine were significantly higher in SB-334867 plus morphine treated rats than that of vehicle plus morphine treated ones on days 4-7. It is concluded that orexin receptor type-1 might be involved in the development of tolerance to morphine analgesic effects.     

Food-induced expression of orexin receptors in rat duodenal mucosa regulates the bicarbonate secretory response to orexin-A

Presence of appetite-regulating peptides orexin-A and orexin-B in mucosal endocrine cells suggests a role in physiological control of the intestine. Our aim was to characterize orexin-induced stimulation of duodenal bicarbonate secretion and modulation of secretory responses and mucosal orexin receptors by overnight food deprivation. Lewis x Dark Agouti rats were anesthetized and proximal duodenum cannulated in situ. Mucosal bicarbonate secretion (pH stat) and mean arterial blood pressure were continuously recorded. Orexin-A was administered intra-arterially close to the duodenum, intraluminally, or into the brain ventricles. Total RNA was extracted from mucosal specimens, reverse transcribed to cDNA and expression of orexin receptors 1 and 2 (OX1 and OX2) measured by quantitative real-time PCR. OX1 protein was measured by Western blot. Intra-arterial orexin-A (60-600 nmol.h(-1).kg(-1)) increased (P < 0.01) the duodenal secretion in fed but not in fasted animals. The OX1 receptor antagonist SB-334867, which was also found to have a partial agonist action, abolished the orexin-induced secretory response but did not affect secretion induced by the muscarinic agonist bethanechol. Atropine, in contrast, inhibited bethanechol but not orexin-induced secretion. Orexin-A infused into the brain ventricles (2-20 nmol.kg(-1).h(-1)) or added to luminal perfusate (1.0-100 nM) did not affect secretion, indicating that orexin-A acts peripherally and at basolateral receptors. Overnight fasting decreased mucosal OX1 and OX2 mRNA expression (P < 0.01) as well as OX1 protein expression (P < 0.05). We conclude that stimulation of secretion by orexin-A may involve both receptor types and is independent of cholinergic pathways. Intestinal OX receptors and secretory responses are markedly related to food intake.     

Dual hypocretin receptor antagonism is more effective for sleep promotion than antagonism of either receptor alone

The hypocretin (orexin) system is involved in sleep/wake regulation, and antagonists of both hypocretin receptor type 1 (HCRTR1) and/or HCRTR2 are considered to be potential hypnotic medications. It is currently unclear whether blockade of either or both receptors is more effective for promoting sleep with minimal side effects. Accordingly, we compared the properties of selective HCRTR1 (SB-408124 and SB-334867) and HCRTR2 (EMPA) antagonists with that of the dual HCRTR1/R2 antagonist almorexant in the rat. All 4 antagonists bound to their respective receptors with high affinity and selectivity in vitro. Since in vivo pharmacokinetic experiments revealed poor brain penetration for SB-408124, SB-334867 was selected for subsequent in vivo studies. When injected in the mid-active phase, SB-334867 produced small increases in rapid-eye-movement (REM) and non-REM (NR) sleep. EMPA produced a significant increase in NR only at the highest dose studied. In contrast, almorexant decreased NR latency and increased both NR and REM proportionally throughout the subsequent 6 h without rebound wakefulness. The increased NR was due to a greater number of NR bouts; NR bout duration was unchanged. At the highest dose tested (100 mg/kg), almorexant fragmented sleep architecture by increasing the number of waking and REM bouts. No evidence of cataplexy was observed. HCRTR1 occupancy by almorexant declined 4-6 h post-administration while HCRTR2 occupancy was still elevated after 12 h, revealing a complex relationship between occupancy of HCRT receptors and sleep promotion. We conclude that dual HCRTR1/R2 blockade is more effective in promoting sleep than blockade of either HCRTR alone. In contrast to GABA receptor agonists which induce sleep by generalized inhibition, HCRTR antagonists seem to facilitate sleep by reducing waking "drive".     

Orexins在雌性大鼠条件性蔗糖摄取中的作用研究

目的:探讨Orexins在雌性大鼠的摄取蔗糖的操作反应行为的作用,以及在线索诱导下恢复寻找蔗糖行为中的作用。方法:将雌性SD大鼠分为限制进食组和自由进食组,以固定比率和累进比率训练大鼠自己摄取蔗糖颗粒。通过Ox R1受体拮抗剂SB-334867预处理,观察SB-334867对大鼠按固定比率摄取蔗糖行为和在线索诱导下恢复寻找蔗糖行为的影响。结果:限制进食的大鼠表现出按压有效杠杆次数和获取蔗糖颗粒数显著增多(P0.05),按压无效杠杆次数稍微增多。SB-334867可显著减少限制进食大鼠按压有效杠杆次数(P0.05)。与对照组相比,SB-334867在消退期间可显著增加按压有效杠杆的次数(P0.05);在恢复期间,限制进食大鼠按压有效杠杆的次数显著增多(P0.05)。结论:Orexins系统在大鼠条件刺激诱导摄取蔗糖中可能存在性别差异。     

SB-334867 (an Orexin-1 Receptor Antagonist) Effects on Morphine-Induced Sensitization in Mice—a View on Receptor Mechanisms

The present study focused upon the role of SB-334867, an orexin-1 receptor antagonist, in the acquisition of morphine-induced sensitization to locomotor activity in mice. Behavioral sensitization is an enhanced systemic reaction to the same dose of an addictive substance, which assumingly increases both the desire for the drug and the risk of relapse to addiction. Morphine-induced sensitization in mice was achieved by sporadic doses (five injections every 3 days) of morphine (10 mg/kg, i.p.), while a challenge dose of morphine (10 mg/kg) was injected 7 days later. In order to assess the impact of orexin system blockade on the acquisition of sensitization, SB-334867 was administered before each morphine injection, except the morphine challenge dose. The locomotor activity test was performed on each day of morphine administration. Brain structures (striatum, hippocampus, and prefrontal cortex) were collected after behavioral tests for molecular experiments in which mRNA expression of orexin, dopamine, and adenosine receptors was explored by the qRT-PCR technique. Additionally, the mRNA expression of markers, such as GFAP and Iba-1, was also analyzed by the same technique. SB-334867 inhibited the acquisition of morphine-induced sensitization to locomotor activity of mice. Significant alterations were observed in mRNA expression of orexin, dopamine, and adenosine receptors and in the expression of GFAP and Iba-1, showing a broad range of interactions in the mesolimbic system among orexin, dopamine, adenosine, and glial cells during behavioral sensitization. Summing up, the orexin system may be an effective measure to inhibit morphine-induced behavioral sensitization.     

1,3-Biarylureas as selective non-peptide antagonists of the orexin-1 receptor.

This communication reports SARs for the first orexin-1 receptor antagonist series of 1-aryl-3-quinolin-4-yl and 1-aryl-3-naphthyridin-4-yl ureas. One of these compounds, 31 (SB-334867), has excellent selectivity for the orexin-1 receptor, blood-brain barrier permeability and shows in vivo activity following ip dosing.     

Orexin在隔核对大鼠胃传入信息的调控

目的:研究orexin在隔核对大鼠胃传入信息的调控作用。方法:选取健康成年雄性Wistar大鼠138只(体质量250-300 g),记录神经元放电活动,鉴定隔核胃牵张(GD)敏感性神经元;隔核微量注射orexin-A或orexin-A受体拮抗剂SB334867,观察隔核GD敏感性神经元放电活动变化;隔核微量注射不同浓度的orexin-A,观察大鼠胃运动的变化。结果:隔核微量注射orexin-A的大鼠胃运动幅度和频率显著增加,并呈剂量依赖关系(P0.05-0.01),微量注射SB-334867可完全阻断orexin-A对胃运动的影响。隔核微量注射orexin-A后,有36个GD-E神经元兴奋(P0.01),16个GD-I神经元抑制。Orexin-A受体拮抗剂SB334867可完全阻断orexin-A对GD敏感神经元的作用。结论:隔核注射orexin能促进大鼠胃运动,并影响胃牵张敏感神经元的放电活动。     

下丘脑腹内侧核Orexin-1及其受体对大鼠胃酸分泌的影响及其机制

目的:探讨下丘脑腹内侧核Orexin-1及其受体对大鼠胃酸分泌的影响及其机制。方法:大鼠麻醉后侧脑室及VMH置管,大鼠分组后分别VMH注射orexin-A、[Pro~(34)]-酪酪肽、[c PP1-7、NPY~(19-23)、Ala~(31)、Aib~(32)、Gln~(34)]胰多肽;腹腔注射SB-334867;皮下注射阿托品;侧脑室微量注射GR-231118、CGP-71683。给药结束后使用幽门结扎模型检测大鼠的胃酸分泌。结果:OXA能够促进胃酸分泌,且呈量效依赖关系。腹腔注射SB-334867能够抑制胃酸分泌,且呈量效依赖关系;SB-334867能够抑制orexin-A对胃酸分泌的促进作用;阿托品不但能够抑制胃酸分泌并且还能够完全阻断OXA的促胃酸分泌作用。侧脑室微量注射GR-231118或CGP-71683胃酸及胃液量减少,呈量效依赖关系,并且能够完全阻断OXA的促胃酸分泌作用。VMH内微量注射[cPP~(1-7),NPY~(19-23),Ala~(31),Aib~(32),Gln~(34)]胰多肽胃酸分泌增多,且呈量效依赖关系。结论:Orexin-A能够作用于下丘脑VMH促进胃酸分泌,orexin受体、Y1和Y5受体以及迷走神经系统均参与该过程。     

A selective orexin-1 receptor antagonist reduces food consumption in male and female rats

A variety of evidence implicates the orexins, especially orexin-A, in the regulation of food intake, but it has not been established whether this effect is mediated by the orexin-1 or orexin-2 receptor. In the present study, a selective orexin-1 receptor antagonist, 1-(2-methylbenzoxazol-6-yl)-3-[1,5]naphthyridin-4-yl urea hydrochloride (SB-334867-A), was administered intraperitoneally to rats under various conditions, and food consumption was subsequently measured over 24 h. In male rats, a single dose of SB-334867-A (30 mg/kg, i.p.) given during the light phase reduced both orexin-A-induced food intake (7 nmol, i.c.v.) and feeding stimulated by an overnight fast for 4 h. When given at the start of the dark phase, food consumption was reduced in both male and female rats over 24 h. Daily injections at the start of the dark phase for 3 days reduced natural feeding in male rats over 24 h on days one and three. These findings demonstrate direct inhibition of orexin-A induced food intake with a selective orexin-1 receptor antagonist. Furthermore, the suppression of nocturnal feeding and food intake stimulated by an overnight fast supports other evidence that orexin-A is involved in the regulation of natural feeding and suggests that orexin-1 receptor antagonists could be useful in the treatment of obesity.     

Discovery of potent and stable conformationally constrained analogues of the MCH R1 antagonist SB-568849

A strategy of systematically targeting more rigid analogues of the known MCH R1 receptor antagonist, SB-568849, serendipitously uncovered a binding mode accessible to N-aryl-phthalimide ligands. Optimisation to improve the stability of this compound class led to the discovery of novel N-aryl-quinazolinones, benzotriazinones and thienopyrimidinones as selective ligands with good affinity for human melanin-concentrating hormone receptor 1.     

Contribution of orexin in hypercapnic chemoreflex: evidence from genetic and pharmacological disruption and supplementation studies in mice.

We have previously shown that hypercapnic chemoreflex in prepro-orexin knockout mice (ORX-KO) is attenuated during wake but not sleep periods. In that study, however, hypercapnic stimulation had been chronically applied for 6 h because of technical difficulty in changing the composition of the inspired gas mixture without distorting the animal's vigilance states. In the present study we examined possible involvement of orexin in acute respiratory chemoreflex during wake periods. Ventilation was recorded together with electroencephalography and electromyography before and after intracerebroventricular administration of orexin or an orexin receptor antagonist, SB-334867. A hypercapnic (5 or 10% CO(2)) or hypoxic (15 or 10% O(2)) gas mixture was introduced into the recording chamber for 5 min. Respiratory parameters were analyzed only for quiet wakefulness. When mice breathed normal room air, orexin-A and orexin-B but not vehicle or SB-334867 increased minute ventilation in both ORX-KO and wild-type (WT) mice. As expected, hypercapnic chemoreflex in vehicle-treated ORX- KO mice (0.22 +/- 0.03 mlxmin(-1)xg(-1)x% CO(2)(-1)) was significantly blunted compared with that in WT mice (0.51 +/- 0.05 mlxmin(-1)xg(-1)x% CO(2)(-1)). Supplementation of orexin-A or -B (3 nmol) partially restored the hypercapnic chemoreflex in ORX-KO mice (0.28 +/- 0.03 mlxmin(-1).g(-1)x% CO(2)(-1) for orexin-A and 0.32 +/- 0.04 mlxmin(-1)xg(-1)x% CO(2)(-1) for orexin-B). In addition, injection of SB-334867 (30 nmol) in WT mice decreased the hypercapnic chemoreflex (0.39 +/- 0.04 mlxmin(-1)xg(-1)x% CO(2)(-1)). On the other hand, hypoxic chemoreflex in vehicle-treated ORX-KO and SB-334867-treated WT mice was not different from that in corresponding controls. Our findings suggest that orexin plays a crucial role in CO(2) sensitivity at least during wake periods in mice.