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.     

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.     

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.     

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.     

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.     

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.     

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.     

Orexins/hypocretins in the ob/ob mouse: hypothalamic gene expression, peptide content and metabolic effects

Orexins (forms A and B) belong to a new family of peptides that, as neuropeptide Y (NPY), stimulate food intake when centrally injected. The ob/ob mouse is a well-characterized model of hyperphagia and obesity associated with strong metabolic disturbances and a central dysregulation of peptides involved in the control of feeding. In the present report, we investigated the hypocretin (Hcrt)/orexin (OX) peptide pathway in lean and ob/ob mice. Prepro-Hcrt/OX mRNA expression, measured by in situ hybridization was restricted to the lateral hypothalamus area. It was significantly decreased in ob/ob mice (-18%; p<0.01). When estimated by real time RT-PCR in the whole hypothalamus, this decrease amounted to 65% (p<0.001). Hcrt-1/OX-A peptide concentrations, measured by RIA in microdissected hypothalamic nuclei were high in the lateral hypothalamus (LH) and lower in the arcuate (ARC) and paraventricular nuclei (PVN). In ob/ob mice, OX-A levels were significantly lower than in lean mice in the LH (-34%; p<0.02) and in the PVN (-72%; p<0.005). Acute intracerebroventricular injection of Hcrt-1/OX-A (1-10 nmol) stimulated feeding in lean, but not in ob/ob mice, whereas Hcrt-2/OX-B (1-10 nmol) had the opposite effect. Acute third ventricle (i3vt) injections of Hcrt/OX peptides in ob/ob mice transiently increased their metabolic rate and stimulated lipid substrate utilization. These findings provide direct evidence that Hcrt/OX peptides are down-regulated in the hypothalamus of ob/ob mice, contrary to the NPY system. The present data argues that Hcrt/OX peptides are not primarily responsible for the metabolic syndrome of the ob/ob mice. The diminution in the OX tone might participate in a counterregulatory system necessary to limit the adverse effects of NPY on food intake and body weight.     

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.     

Anti-obesity and Antidiabetic Effects of Deep Sea Water on ob/ob Mice

Recently, deep sea water (DSW) has started to receive much attention for therapeutic intervention in some lifestyle diseases. In this study, the anti-obesity and antidiabetic effects of DSW in ob/ob mice were investigated. The animals were randomly divided into two groups with six animals: control group received tap water; the experimental group was treated with DSW of hardness 1000 for 84 days. The body weight gain after 84 days in DSW-fed group was decreased by 7% compared to the control group. The plasma glucose levels in the DSW-fed mice were substantially reduced by 35.4%, as compared to control mice. The results of oral glucose tolerance test revealed that DSW-fed groups significantly increased the glucose disposal after 84 days. DSW increased plasma protein levels of adiponectin and decreased plasma protein levels of resistin, RBP4, and fatty acid binding protein. Moreover, GLUT4 and AMP-activated protein kinase levels in skeletal muscle tissue were increased while peroxisome proliferator-activated receptor γ and adiponectin were decreased in adipose tissue of DSW-fed mice. These results suggest that the antidiabetic and anti-obesity activities of DSW were mediated by modulating the expression of diabetes- and obesity-specific molecules. Taken together, these results provide a possibility that continuous intake of DSW can ameliorate obesity and diabetes.     

Diaryl urea analogues of SB-334867 as orexin-1 receptor antagonists

As a part of our program to develop OX1-CB1 bivalent ligands, we required a better understanding of the basic structure-activity relationships (SARs) of orexin antagonists. A series of SB-334867 analogues were synthesized and evaluated in calcium mobilization assays. SAR results suggest that the 2-methylbenzoxazole moiety may be replaced with a disubstituted 4-aminophenyl group without loss of activity and an electron-deficient system is generally preferred at the 1,5-naphthyridine moiety for OX1 antagonist activity. In particular, substitution of larger potential linkers such as n-hexyl provided compound 33 with equivalent activity at the OX1 receptor compared to the lead compound SB-334867. These compounds should be of value in the development of ligands targeting the orexin-1 receptor and its potential heterodimers.     

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.     

Effects of leptin replacement on hypothalamic-pituitary growth hormone axis function and circulating ghrelin levels in ob/ob mice

Leptin-deficient obese mice (ob/ob) have decreased circulating growth hormone (GH) and pituitary GH and ghrelin receptor (GHS-R) mRNA levels, whereas hypothalamic GH-releasing hormone (GHRH) and somatostatin (SST) expression do not differ from lean controls. Given the fact that GH is suppressed in diet-induced obesity (a state of hyperleptinemia), it remains to be determined whether the absence of leptin contributes to changes in the GH axis of ob/ob mice. Therefore, to study the impact of leptin replacement on the hypothalamic-pituitary GH axis of ob/ob mice, leptin was infused for 7 days (sc), resulting in circulating leptin levels that were similar to wild-type controls (approximately 1 ng/ml). Leptin treatment reduced food intake, body weight, and circulating insulin while elevating circulating n-octanoyl ghrelin concentrations. Leptin treatment did not alter hypothalamic GHRH, SST, or GHS-R mRNA levels compared with vehicle-treated controls. However, leptin significantly increased pituitary GH and GHRH-R expression and tended to enhance circulating GH levels, but this latter effect did not reach statistical significance. In vitro, leptin (1 ng/ml, 24 h) did not affect pituitary GH, GHRH-R, or GHS-R mRNA but did enhance GH release. The in vivo effects of leptin on circulating hormone and pituitary mRNA levels were not replicated by pair feeding ob/ob mice to match the food intake of leptin-treated mice. However, leptin did prevent the fall in hypothalamic GHRH mRNA and circulating IGF-I levels observed in pair-fed mice. These results demonstrate that leptin replacement has positive effects on multiple levels of GH axis function in ob/ob mice.     

Adrenalectomy in genetically obese ob/ob and db/db mice increases the proton conductance pathway

Adrenalectomy (ADX) prevents the excessive weight gain in the genetically obese ob/ob and db/db mice. To test the possibility that this results from increased energy expenditure due to increased thermogenesis in brown adipose tissue (BAT), we measured GDP binding to mitochondria from interscapular brown adipose tissue (BAT) in db/db and ob/ob mice and their lean controls after adrenalectomy, with and without corticosterone replacement. Both the vehicle treated and corticosterone treated db/db and ob/ob mice had lower body weights than the sham-operated mice GDP binding to mitochondria from IBAT was significantly lower in both the db/db and ob/ob mice than in their lean controls. Adrenalectomy significantly increased GDP binding in all mice compared to the respective sham-operated mice, but, the percentage increase was always greater in the db/db and ob/ob mice. Corticosterone treatment of adrenalectomized db/db, ob/ob or lean mice lowered GDP binding to sham levels. Our data confirm previous findings that adrenalectomy results in increased GDP binding to mitochondria from IBAT. Injections of corticosterone into adrenalectomized mice results in a decrease in GDP binding to values which are similar to values in sham-operated mice. Thus adrenalectomy may inhibit the development of obesity by increasing the thermic activity in IBAT.     

A Relationship between Reduced Nucleus Accumbens Shell and Enhanced Lateral Hypothalamic Orexin Neuronal Activation in Long-Term Fructose Bingeing Behavior

Fructose accounts for 10% of daily calories in the American diet. Fructose, but not glucose, given intracerebroventricularly stimulates homeostatic feeding mechanisms within the hypothalamus; however, little is known about how fructose affects hedonic feeding centers. Repeated ingestion of sucrose, a disaccharide of fructose and glucose, increases neuronal activity in hedonic centers, the nucleus accumbens (NAc) shell and core, but not the hypothalamus. Rats given glucose in the intermittent access model (IAM) display signatures of hedonic feeding including bingeing and altered DA receptor (R) numbers within the NAc. Here we examined whether substituting fructose for glucose in this IAM produces bingeing behavior, alters DA Rs and activates hedonic and homeostatic feeding centers. Following long-term (21-day) exposure to the IAM, rats given 8–12% fructose solutions displayed fructose bingeing but unaltered DA D1R or D2R number. Fructose bingeing rats, as compared to chow bingeing controls, exhibited reduced NAc shell neuron activation, as determined by c-Fos-immunoreactivity (Fos-IR). This activation was negatively correlated with orexin (Orx) neuron activation in the lateral hypothalamus/perifornical area (LH/PeF), a brain region linking homeostatic to hedonic feeding centers. Following short-term (2-day) access to the IAM, rats exhibited bingeing but unchanged Fos-IR, suggesting only long-term fructose bingeing increases Orx release. In long-term fructose bingeing rats, pretreatment with the Ox1R antagonist SB-334867 (30 mg/kg; i.p.) equally reduced fructose bingeing and chow intake, resulting in a 50% reduction in calories. Similarly, in control rats, SB-334867 reduced chow/caloric intake by 60%. Thus, in the IAM, Ox1Rs appear to regulate feeding based on caloric content rather than palatability. Overall, our results, in combination with the literature, suggest individual monosaccharides activate distinct neuronal circuits to promote feeding behavior. Specifically, long-term fructose bingeing activates a hyperphagic circuit composed in part of NAc shell and LH/PeF Orx neurons.     

Behavioral characteristics and pharmacological manipulations of a nicotine-entrainable circadian oscillator

Chronic daily administration of nicotine and other drugs of abuse has been found to entrain pre- and post-drug circadian locomotor activity episodes that oscillate on a 24-h schedule and persist for several days after administration ceases. This drug-entrainable oscillator system could conceivably lead to circadian rhythms of drug seeking and drug use in human drug addicts. The present study (1) characterizes the ability of daily nicotine administration to entrain circadian wheel-running activity episodes in rats across a range of doses, lighting schedules, and food access; and (2) tests whether pre- and post-nicotine episodes can be altered through pharmacological manipulations. Adult female rats were housed in wheel boxes for 35–60?d, and both wheel-running and feeding-related behaviors were measured continuously. Following acclimation, nicotine or saline was administered for 16–24?d, and the rats were left undisturbed for several test days to observe the persistence of nicotine-entrained activity. The results showed that nicotine dose-dependently entrains wheel-running activity, and the highest dose of 1.0?mg/kg produces robust pre- and post-nicotine circadian activity episodes under constant, fixed, and variable light/dark schedules. In the pharmacological manipulation experiment, nicotine-entrained rats were administered one of seven pharmacological treatments (varenicline, mecamylamine, acamprosate, topiramate, naltrexone, SB-334867, or bupropion) in place of the nicotine injection for 2?d, and the rats were not disturbed for four subsequent days. Most of the treatment drugs significantly reduced post-nicotine activity episodes, but only three treatments affected pre-nicotine episodes: the μ- and κ-opioid receptor antagonist naltrexone, the orexin-1 receptor antagonist SB-334867, and the AMPA (2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid)/kainate antagonist topiramate. These results show that chronic daily nicotine administration is a robust zeitgeber that dose-dependently entrains a nonphotic oscillator system that includes opioid, orexin, and glutamate pathways.     

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.     

Effect of clofibrate on malic enzyme and leptin mRNAs level in rat brown and white adipose tissue.

Two previous studies have reported contradictory results regarding the effect of fibrates treatment on obese (ob) gene expression in rodents. The purpose of the present study was to reinvestigate this issue. We examined the effect of clofibrate (fibrate derivative) administration for 14 days to rats on malic enzyme (as an adequate control of fibrates action) and leptin mRNAs level in the white and brown adipose tissues (WAT and BAT, respectively). The malic enzyme activity and malic enzyme mRNA level in white adipose tissue increased significantly after clofibrate feeding. In brown adipose tissue, the drug treatment resulted in depression of malic enzyme activity and malic enzyme mRNA level. Under the same conditions, leptin mRNA level did not change in these tissues. The results presented in this paper provide further evidence that the clofibrate (activator of peroxisome proliferator activated receptor alpha), feeding is without effect on ob gene expression in rat white and brown adipose tissue. Furthermore, the present study demonstrates that clofibrate causes opposite effects on malic enzyme gene expression in WAT (up-regulation) and BAT (down-regulation).