Discovery of a benzimidazole series as the first highly potent and selective ACSL1 inhibitors.

Long-chain acyl-CoA synthetase-1 (ACSL1), an enzyme that catalyzes the synthesis of long-chain acyl-CoA from the corresponding fatty acids, is believed to play essential roles in lipid metabolism. Structure activity relationship studies based on HTS hit compound 1 delivered the benzimidazole series as the first selective and highly potent ACSL1 inhibitors. Representative compound 13 exhibited not only remarkable inhibitory activity against ACSL1 (IC50 = 0.042 µM) but also excellent selectivity for the other ACSL isoforms. In addition, compound 13 demonstrated an in vivo suppression effect against the production of long-chain acyl-CoAs in mouse.

Promotion of crystalline cellulose degradation by expansins from Oryza sativa.

MAIN CONCLUSION: Enzymatic activities of Oryza sativa expansins, which were heterologously overexpressed in Escherichia coli , were analyzed. Results suggested that expansins promote degradation of cellulose by cellulase in a synergistic manner. Sustainable production of future biofuels is dependent on efficient saccharification of lignocelluloses. Expansins have received a lot of attention as proteins promoting biological degradation of cellulose using cellulase. The expansins are a class of plant cell wall proteins that induce cell wall loosening without hydrolysis. In this study, the expansins from Oryza sativa were classified using phylogenetic analysis and five proteins were selected for functional evaluation. At low cellulose loading, the cellulase in expansin mixtures was up to 2.4 times more active than in mixtures containing only cellulase, but at high cellulose loading the activity of cellulase in expansin mixtures and cellulase only mixtures did not differ. Furthermore, expansin activity was greater in cellulase mixtures compared with cellulase-deficient mixtures. Therefore, the expansins showed significant synergistic activity with cellulase. Expansin may play an important role in efficient saccharification of cellulose.

Phase 1 and pharmacological trial of OPB-31121, a signal transducer and activator of transcription-3 inhibitor, in patients with advanced hepatocellular carcinoma.

AIM: To evaluate the safety, pharmacokinetics and antitumor activity of OPB-31121, a signal transducer and activator of transcription-3 inhibitor, in patients with advanced hepatocellular carcinoma (HCC). METHODS: HCC patients of Child-Pugh A or B who progressed on, or were intolerant to, sorafenib were eligible for this phase 1 trial. We used a standard 3 + 3 dose-escalation design with a 28-day cycle at dose levels of 50, 100, 200 and 400 mg/day. Tumor responses were assessed using the modified Response Evaluation Criteria in Solid Tumors. RESULTS: Twenty-four patients were enrolled, of whom 23 received OPB-31121 (20 males; median age, 65 years). The most common adverse drug reactions were nausea (87.0%), vomiting (82.6%), diarrhea (69.6%), fatigue/malaise (52.2%), anorexia (47.8%) and peripheral sensory neuropathy (26.1%). The recommended dose for OPB-31121 was determined to be 200 mg. Six patients had stable disease for 8 weeks or more, resulting in disease control rates of 25.0-42.9%. In the 200-mg dose cohort, three of seven patients had stable disease and a median time to progression of 61.0 days. The maximum concentration and area under the plasma concentration-time curve of OPB-31121 were dose proportional. CONCLUSION: OPB-31121 demonstrated insufficient antitumor activity for HCC. Furthermore, peripheral nervous system-related toxicities may negatively affect long-term administration of OPB-31121. Therefore, it was deemed difficult to continue the clinical development of OPB-31121 for treating advanced HCC and further investigation is expected in the agent with favorable profile in this category.

Acsl1 is essential for skin barrier function through the activation of linoleic acid and biosynthesis of ω-O-acylceramide in mice.

The long-chain acyl-CoA synthase1 (Acsl1) is a major enzyme that converts long-chain fatty acids to acyl-CoAs. The role of Acsl1 in energy metabolism has been elucidated in the adipose tissue, heart, and skeletal muscle. Here, we demonstrate that systemic deficiency of Acsl1 caused severe skin barrier defects, leading to embryonic lethality. Acsl1 mRNA and protein are expressed in the Acsl1+/+ epidermis, which are absent in Acsl1-/- mice. In Acsl1-/- mice, epidermal ceramide [EOS] (Cer[EOS]) containing ω-O-esterified linoleic acid, a lipid essential for the skin barrier, was significantly reduced. Conversely, ω-hydroxy ceramide (Cer[OS]), a precursor of Cer[EOS], was increased. Moreover, the levels of triglyceride (TG) species containing linoleic acids were lower in Acsl1-/- mice, whereas those not containing linoleic acid were comparable to Acsl1+/+ mice. As TG is considered to work as a reservoir of linoleic acid for the biosynthesis of Cer[EOS] from Cer[OS], our results suggest that Acsl1 plays an essential role in ω-O-acylceramide synthesis by providing linoleic acid for ω-O-esterification. Therefore, our findings identified a new biological role of Acsl1 as a regulator of the skin barrier.

Lectin-like oxidized LDL receptor 1 is involved in CRP-mediated complement activation.

BACKGROUND: C-reactive protein (CRP) is purported to be a risk factor that acts independently of LDL cholesterol in predicting all-cause mortality in patients with ischemic heart disease. Lectin-like oxidized LDL receptor 1 (LOX-1) impairs endothelial function and exacerbates myocardial injury. We recently demonstrated that CRP increased vascular permeability through direct binding to LOX-1. Here we examined, using a hypertensive rat model, whether LOX-1 is involved in CRP-induced complement activation. METHODS AND RESULTS: In the cultured LOX-1-expressing cell line hLOX-1-CHO, CRP increased complement activation, but did not do so in native CHO cells. Depleting C1q from serum abolished CRP-induced complement activation. Incubation of CRP with serum on immobilized recombinant LOX-1 similarly showed that CRP activated C1q-requiring classical complement pathway in a LOX-1-dependent manner. Interestingly, the interaction between CRP and LOX-1 was dependent on Ca²âº ion and competed with phosphocholine, suggesting that LOX-1 bound to the B-face of CRP with a phosphocholine-binding domain. This was in contrast to Fcγ receptors, to which CRP bound in A-face with complement-binding domain. In vivo, intradermal injection of CRP to hypertensive SHRSP rats induced complement activation detected by C3d deposition and leukocyte infiltration around the injected area. Anti-LOX-1 antibody reduced the extent of complement activation and leukocyte infiltration. CONCLUSIONS: LOX-1 appears to be involved in CRP-induced complement activation, and thus may serve to locate the site of CRP-induced complement activation and inflammation.

The discovery of LOX-1, its ligands and clinical significance.

LOX-1 is an endothelial receptor for oxidized low-density lipoprotein (oxLDL), a key molecule in the pathogenesis of atherosclerosis.The basal expression of LOX-1 is low but highly induced under the influence of proinflammatory and prooxidative stimuli in vascular endothelial cells, smooth muscle cells, macrophages, platelets and cardiomyocytes. Multiple lines of in vitro and in vivo studies have provided compelling evidence that LOX-1 promotes endothelial dysfunction and atherogenesis induced by oxLDL. The roles of LOX-1 in the development of atherosclerosis, however, are not simple as it had been considered. Evidence has been accumulating that LOX-1 recognizes not only oxLDL but other atherogenic lipoproteins, platelets, leukocytes and CRP. As results, LOX-1 not only mediates endothelial dysfunction but contributes to atherosclerotic plaque formation, thrombogenesis, leukocyte infiltration and myocardial infarction, which determine mortality and morbidity from atherosclerosis. Moreover, our recent epidemiological study has highlighted the involvement of LOX-1 in human cardiovascular diseases. Further understandings of LOX-1 and its ligands as well as its versatile functions will direct us to ways to find novel diagnostic and therapeutic approaches to cardiovascular disease.

Procyanidins are potent inhibitors of LOX-1: a new player in the French Paradox.

Lectin-like oxidized LDL receptor-1 (LOX-1) is an endothelial receptor for oxidized LDL (oxLDL) and plays multiple roles in the development of cardiovascular diseases. We screened more than 400 foodstuff extracts for identifying materials that inhibit oxLDL binding to LOX-1. Results showed that 52 extracts inhibited LOX-1 by more than 70% in cell-free assays. Subsequent cell-based assays revealed that a variety of foodstuffs known to be rich in procyanidins such as grape seed extracts and apple polyphenols, potently inhibited oxLDL uptake in Chinese hamster ovary (CHO) cells expressing LOX-1. Indeed, purified procyanidins significantly inhibited oxLDL binding to LOX-1 while other ingredients of apple polyphenols did not. Moreover, chronic administration of oligomeric procyanidins suppressed lipid accumulation in vascular wall in hypertensive rats fed with high fat diet. These results suggest that procyanidins are LOX-1 inhibitors and LOX-1 inhibition might be a possible underlying mechanism of the well-known vascular protective effects of red wine, the French Paradox.

Electric dipole induced bulk ferromagnetism in dimer Mott molecular compounds.

Magnetic properties of Mott-Hubbard systems are generally dominated by strong antiferromagnetic interactions produced by the Coulomb repulsion of electrons. Although theoretical possibility of a ferromagnetic ground state has been suggested by Nagaoka and Penn as single-hole doping in a Mott insulator, experimental realization has not been reported more than half century. We report the first experimental possibility of such ferromagnetism in a molecular Mott insulator with an extremely light and homogeneous hole-doping in π-electron layers induced by net polarization of counterions. A series of Ni(dmit)2 anion radical salts with organic cations, where dmit is 1,3-dithiole-2-thione-4,5-dithiolate can form bi-layer structure with polarized cation layers. Heat capacity, magnetization, and ESR measurements substantiated the formation of a bulk ferromagnetic state around 1.0 K with quite soft magnetization versus magnetic field (M-H) characteristics in (Et-4BrT)[Ni(dmit)2]2 where Et-4BrT is ethyl-4-bromothiazolium. The variation of the magnitude of net polarizations by using the difference of counter cations revealed the systematic change of the ground state from antiferromagnetic one to ferromagnetic one. We also report emergence of metallic states through further doping and applying external pressures for this doping induced ferromagnetic state. The realization of ferromagnetic state in Nagaoka-Penn mechanism can paves a way for designing new molecules-based ferromagnets in future.

In vivo imaging of obesity-induced inflammation in adipose tissue.

Obesity is associated with low-grade inflammation in adipose tissue, which contributes to the development of obesity-related diseases such as insulin resistance, hypertension and arteriosclerosis. Here we developed an animal model that non-invasively monitors inflammation in adipose tissue using in vivo bioluminescent imaging (BLI) technique. In vitro, stimulation with TNFalpha or co-culture with RAW264 macrophages increased bioluminescence in 3T3-L1 adipocytes expressing NF-kappaB-mediated luciferase gene (3T3-L1/NF-kappaB-re-luc2P). In vivo, lipopolysaccharide increased bioluminescence in mice transplanted with 3T3-L1/NF-kappaB-re-luc2P cells. Moreover, light emission derived from implanted cells was significantly higher in diet-induced obese mice transplanted with 3T3-L1/NF-kappaB-re-luc2P than in lean mice. Our results showed that BLI technique and 3T3-L1/NF-kappaB-re-luc2P cells provide a useful approach to non-invasively monitor obesity-induced inflammation in adipose tissue in in vivo.

C-reactive protein uptake by macrophage cell line via class-A scavenger receptor.

BACKGROUND: C-reactive protein (CRP) increases in response to inflammation and is purported to be a risk factor for atherogenesis. We recently demonstrated that a scavenger receptor, lectin-like oxidized LDL receptor (LOX-1), is a receptor for CRP. In light of the overlapping ligand spectrum of scavenger receptors such as modified LDL, bacteria, and advanced glycation end products, we examined whether other scavenger receptors recognize CRP. METHODS: We analyzed the uptake of fluorescently labeled CRP in COS-7 cells expressing a series of scavenger receptors and in a monocytic cell line, THP-1, differentiated into macrophage with phorbol 12-myristate 13-acetate (PMA). We applied small interfering RNA (siRNA) against class-A scavenger receptor (SR-A) to THP-1 cells to suppress the expression of SR-A. We also analyzed the binding of nonlabeled CRP to immobilized recombinant LOX-1 and SR-A in vitro using anti-CRP antibody. RESULTS: COS-7 cells expressing LOX-1 and SR-A internalized fluorescently labeled CRP in a dose-dependent manner, but cells expressing CD36, SR-BI, or CD68 did not. The recombinant LOX-1 and SR-A proteins recognized nonlabeled purified CRP and native CRP in serum in vitro. THP-1 cells differentiated into macrophage-like cells by treatment with PMA-internalized fluorescently labeled CRP. siRNA against SR-A significantly and concomitantly inhibited the expression of SR-A (P < 0.01) and CRP uptake (P < 0.01), whereas control siRNA did not. CONCLUSIONS: CRP is recognized by SR-A as well as LOX-1 and taken up via SR-A in a macrophage-like cell line. This process might be of significance in the pathogenesis of atherosclerotic disease.

LOX index, a novel predictive biochemical marker for coronary heart disease and stroke.

BACKGROUND: Lectin-like oxidized LDL receptor 1 (LOX-1) is implicated in atherothrombotic diseases. Activation of LOX-1 in humans can be evaluated by use of the LOX index, obtained by multiplying the circulating concentration of LOX-1 ligands containing apolipoprotein B (LAB) times that of the soluble form of LOX-1 (sLOX-1) [LOX index = LAB x sLOX-1]. This study aimed to establish the prognostic value of the LOX index for coronary heart disease (CHD) and stroke in a community-based cohort. METHODS: An 11-year cohort study of 2437 residents age 30-79 years was performed in an urban area located in Japan. Of these, we included in the analysis 1094 men and 1201 women without history of stroke and CHD. We measured LAB and sLOX-1 using ELISAs with recombinant LOX-1 and monoclonal anti-apolipoprotein B antibody and with 2 monoclonal antibodies against LOX-1, respectively. RESULTS: During the follow-up period, there were 68 incident cases of CHD and 91 cases of stroke (with 60 ischemic strokes). Compared with the bottom quartile, the hazard ratio (HR) of the top quartile of LOX index was 1.74 (95% CI 0.92-3.30) for stroke and 2.09 (1.00-4.35) for CHD after adjusting for sex, age, body mass index, drinking, smoking, hypertension, diabetes, non-HDL cholesterol, and use of lipid-lowering agents. Compared with the bottom quartile of LOX index, the fully adjusted HRs for ischemic stroke were consistently high from the second to the top quartile: 3.39 (95% CI 1.34-8.53), 3.15 (1.22-8.13) and 3.23 (1.24-8.37), respectively. CONCLUSIONS: Higher LOX index values were associated with an increased risk of CHD. Low LOX index values may be protective against ischemic stroke.

Diacylglycerol acyltransferase 1/2 inhibition induces dysregulation of fatty acid metabolism and leads to intestinal barrier failure and diarrhea in mice.

The intestinal metabolism and transport of triacylglycerol (TAG) play a critical role in dietary TAG absorption, and defects in the process are associated with congenital diarrhea. The final reaction in TAG synthesis is catalyzed by diacylglycerol acyltransferase (DGAT1 and DGAT2), which uses activated fatty acids (FA) as substrates. Loss-of-function mutations in DGAT1 cause watery diarrhea in humans, but mechanisms underlying the relationship between altered DGAT activity and diarrhea remain largely unclear. Here, the effects of DGAT1 and DGAT2 inhibition, alone or in combination, on dietary TAG absorption and diarrhea in mice were investigated by using a selective DGAT1 inhibitor (PF-04620110) and DGAT2 inhibitor (PF-06424439). Simultaneous administration of a single dosing of these inhibitors drastically decreased intestinal TAG secretion into the blood circulatory system and TAG accumulation in the duodenum at 60 min after lipid gavage. Under 60% high-fat diet (HFD) feeding, their repeated simultaneous administration for 2 days induced severe watery diarrhea and occasionally led to death. The diarrhea was accompanied by enhanced fecal FA excretion, intestinal injury and barrier failure. DGAT1 or DGAT2 inhibition alone did not induce the phenotypic changes observed in DGAT1/2 inhibitor-treated mice. The results demonstrate that DGAT1/2 inhibition alters TAG absorption and results in watery diarrhea in mice. DGAT1/2 inhibition-induced diarrhea may be caused by intestinal barrier dysfunction due to dysregulation of the cytotoxic FA metabolism. These findings suggest that DGAT-mediated intestinal TAG synthesis is a vital step for maintaining intestinal barrier integrity under HFD feeding.

Role of MGAT2 and DGAT1 in the release of gut peptides after triglyceride ingestion.

Triglyceride ingestion releases gut peptides from enteroendocrine cells located in the intestinal epithelia and provides feedback regulations of gastrointestinal function. The precise mechanisms sensing lipids in the intestinal wall, however, are not well characterized. In the current study, we investigated the release of gut peptides following oral triglyceride loading in mice deficient for monoacylglycerol acyltransferase 2 (MGAT2KO) and diacylglycerol acyltransferase 1 (DGAT1KO), enzymes that sequentially re-synthesize triglyceride to secrete as chylomicron at the small intestine. In wild-type (Wt) mice, oral triglyceride loading resulted in hypertriglycemia. In addition, plasma glucose-dependent insulinotropic polypeptide (GIP), glucagon-like peptide-1 (GLP-1) and peptide YY (PYY) were significantly increased 30 min after triglyceride loading, before decaying in 2h. In MGAT2KO and DGAT1KO mice, oral triglyceride loading did not result in hypertriglycemia and the increase in GIP was significantly suppressed in both KO mouse strains. In contrast, the increases in plasma GLP-1 and PYY in both KO mouse strains were comparable to Wt mice 30 min after triglyceride loading, however, they remained elevated in DGAT1KO mice even 2h after triglyceride loading. In parallel to the changes in GLP-1 and PYY, gastric emptying was delayed after oral triglyceride loading in MGAT2KO mice comparably to Wt type mice and was further delayed in DGAT1KO mice. STC-1 and GLUTag, GLP-1-producing intestinal endocrine L-cell lines, displayed a significant level of DGAT1 activity but not MGAT activity. These findings suggest that synthesis and/or secretion of triglyceride-rich lipoproteins play an important role in the release of GIP. Moreover, DGAT1 may directly regulate the release of GLP-1 and PYY in L-cells.

5,5-Dimethyl-3-(5-methyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)-1-phenyl-3-(trifluoromethyl)-3,5,6,7-tetrahydro-1H-indole-2,4-dione, a potent inhibitor for mammalian elongase of long-chain fatty acids family 6: examination of its potential utility as a pharmacological tool.

Long-chain fatty acid elongases reside in the endoplasmic reticulum and are responsible for the rate-limiting step of the elongation of long-chain fatty acids. The elongase of long-chain fatty acids (ELOVL) family 6 (ELOVL6) is involved in the elongation of saturated and monosaturated fatty acids. Increased expression of ELOVL6 in ob/ob mice suggests a role for ELOVL6 in metabolic disorders. Furthermore, ELOVL6-deficient mice are protected from high-fat diet-induced insulin resistance, which suggests that ELOVL6 might be a new therapeutic target for diabetes. As reported previously, we developed a high-throughput screening system for fatty acid elongases and discovered lead chemicals that possess inhibitory activities against ELOVL6. In the present study, we examined in detail the biochemical and pharmacological properties of 5,5-dimethyl-3-(5-methyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)-1-phenyl-3-(trifluoromethyl)-3,5,6,7-tetrahydro-1H-indole-2,4-dione (Compound-A), a potent inhibitor of ELOVL6. In in vitro assays, Compound-A dose-dependently inhibited mouse and human ELOVL6 and displayed more than 30-fold greater selectivity for ELOVL6 over the other ELOVL family members. In addition, Compound-A effectively reduced the elongation index of fatty acids of hepatocytes, suggesting that Compound-A penetrates the cell wall and inhibits ELOVL6. More importantly, upon oral administration to mice, Compound-A showed high plasma and liver exposure and potently reduced the elongation index of the fatty acids of the liver. This is the first study to report a potent and selective inhibitor of mammalian elongases. Furthermore, Compound-A seems to be a useful tool to further understand the physiological roles of ELOVL6 and to evaluate the therapeutic potential of an ELOVL6 inhibitor.

Synthesis, structure-activity relationships, and biological profiles of a dihydrobenzoxathiin class of histamine H(3) receptor inverse agonists.

A series of novel dihydrobenzoxathiin derivatives was synthesized and evaluated as potent human histamine H(3) receptor inverse agonists. After systematic modification of lead 1a, the potent and selective histamine H(3) inverse agonist 1-(3-{4-[(2S,3S)-8-methoxy-3-methyl-4,4-dioxido-2,3-dihydro-1,4-benzoxathiin-2-yl]phenoxy}propyl)pyrrolidine (5k) was identified. Compound 5k showed good pharmacokinetic profiles and brain penetrability in laboratory animals. After 3mg/kg oral administration of 5k, significant elevation of brain histamine levels was observed in rats where the brain H(3) receptor was fully occupied.

Lox-1: the multifunctional receptor underlying cardiovascular dysfunction.

Oxidatively modified low-density lipoprotein (oxLDL) is implicated in the pathogenesis of atherosclerosis. Endothelial dysfunction is the initial change in the vascular wall that induces morphological changes for atheroma-formation. Lectin-like oxidized LDL receptor-1 (LOX-1) was identified as the receptor for oxLDL that was thought to be a major cause of endothelial dysfunction. LOX-1 has been demonstrated to contribute not only to endothelial dysfunction, but also to atherosclerotic-plaque formation, myocardial infarction and intimal thickening after balloon injury. Recent findings on the genetics of LOX-1 and the methodology to detect it and its ligands would further facilitate the examination of the receptor's pathophysiological contribution in atherosclerosis. Furthermore, LOX-1-related tools might open new gateways from diagnosis to therapeutics for cardiovascular diseases.

Synthesis and evaluation of a novel 2-azabicyclo[2.2.2]octane class of long chain fatty acid elongase 6 (ELOVL6) inhibitors.

A series of novel 2-azabicyclo[2.2.2]octane derivatives was synthesized and evaluated as long chain fatty acid elongase 6 (ELOVL6) inhibitors. Screening of our corporate chemical collections against ELOVL6 resulted in the identification of lead 1. Exploratory chemistry efforts were applied to lead 1 to identify the orally available, potent, and selective ELOVL6 inhibitor 28a.

Artificial intelligence predicts the progression of diabetic kidney disease using big data machine learning.

Artificial intelligence (AI) is expected to support clinical judgement in medicine. We constructed a new predictive model for diabetic kidney diseases (DKD) using AI, processing natural language and longitudinal data with big data machine learning, based on the electronic medical records (EMR) of 64,059 diabetes patients. AI extracted raw features from the previous 6 months as the reference period and selected 24 factors to find time series patterns relating to 6-month DKD aggravation, using a convolutional autoencoder. AI constructed the predictive model with 3,073 features, including time series data using logistic regression analysis. AI could predict DKD aggravation with 71% accuracy. Furthermore, the group with DKD aggravation had a significantly higher incidence of hemodialysis than the non-aggravation group, over 10 years (N = 2,900). The new predictive model by AI could detect progression of DKD and may contribute to more effective and accurate intervention to reduce hemodialysis.

Low-carbohydrate diet by staple change attenuates postprandial GIP and CPR levels in type 2 diabetes patients.

AIMS: The aim of this study is to investigate the effects of a low-carbohydrate staple food (i.e., low-carbohydrate bread) on glucose and lipid metabolism and pancreatic and enteroendocrine hormone secretion in comparison with meals containing normal-carbohydrate bread, without consideration of the carbohydrate content of the side dishes. METHODS: T2DM patients (n = 41) were provided meals containing low-carbohydrate bread (LB) together with side dishes or normal-carbohydrate bread (NB) together with side dishes every other day as a breakfast. Blood glucose levels were evaluated by using a continuous glucose monitoring system; blood samples were collected before and 1 and 2 h after the breakfast. RESULTS: Postprandial blood glucose levels, plasma insulin, plasma glucose-dependent insulinotropic polypeptide (GIP) and plasma triglyceride were significantly lower and plasma glucagon levels were significantly higher in LB compared with those in NB. Plasma glucagon-like peptide-1 (GLP-1) levels did not differ in the LB and NB groups. CONCLUSIONS: These results indicate that changing only the carbohydrate content of the staple food has benefits on glucose and lipid metabolism in T2DM patients concomitant with the decrease of insulin and GIP secretion, which ameliorate body weight gain and insulin resistance.

Distinctive role of central histamine H3 receptor in various orexigenic pathways.

Despite the well-established role of histamine as an anorexigenic neurotransmitter, the role of histamine H(3) receptors in feeding behavior is controversial. Herein we investigated the role of histamine H(3) receptor on several orexigenic agents in mice. Thioperamide (histamine H(3) receptor inverse agonist) inhibited neuropeptide Y- and nociceptin-induced hyperphagia but had no effect on U-50488 (opioid kappa-receptor agonist)-induced hyperphagia. In contrast, imetit (histamine H(3) receptor agonist) inhibited U-50488-induced hyperphagia but augmented neuropeptide Y-induced hyperphagia while it did not alter nociceptin-induced hyperphagia. These results indicate distinctive roles of histamine H(3) receptors in various orexigenic pathways.