A specific G9a inhibitor unveils BGLT3 lncRNA as a universal mediator of chemically induced fetal globin gene expression.

Sickle cell disease (SCD) is a heritable disorder caused by ß-globin gene mutations. Induction of fetal γ-globin is an established therapeutic strategy. Recently, epigenetic modulators, including G9a inhibitors, have been proposed as therapeutic agents. However, the molecular mechanisms whereby these small molecules reactivate γ-globin remain unclear. Here we report the development of a highly selective and non-genotoxic G9a inhibitor, RK-701. RK-701 treatment induces fetal globin expression both in human erythroid cells and in mice. Using RK-701, we find that BGLT3 long non-coding RNA plays an essential role in γ-globin induction. RK-701 selectively upregulates BGLT3 by inhibiting the recruitment of two major γ-globin repressors in complex with G9a onto the BGLT3 gene locus through CHD4, a component of the NuRD complex. Remarkably, BGLT3 is indispensable for γ-globin induction by not only RK-701 but also hydroxyurea and other inducers. The universal role of BGLT3 in γ-globin induction suggests its importance in SCD treatment.

Dynamics of Oxidative Stress Evoked by Myocardial Ischemia Reperfusion After Off-Pump Coronary Artery Bypass Grafting Elucidated by Bilirubin Oxidation.

BACKGROUND: Revascularization therapy relieves myocardial ischemia, but can also result in ischemia-reperfusion injury caused by oxidative stress. However, the biokinetics of oxidative stress after myocardial ischemia-reperfusion are uncertain. This study aimed to evaluate the dynamics of oxidative stress after off-pump coronary artery bypass grafting (OPCAB) by measuring urinary biopyrrin levels. Biopyrrin is an oxidative metabolite of bilirubin thought to reflect oxidative stress, along with reactive nitrogen species (RNS).Methods and Results:The study included 18 patients who underwent OPCAB; patients were divided into effort angina pectoris (EAP; n=11) and unstable angina pectoris (UAP; n=7). Urinary biopyrrin and RNS levels were measured during the perioperative period (≤48 h after surgery). Biopyrrin levels transiently increased 4-12 h post-surgery (early phase), followed by a prolonged increase approximately 24-32 h post-surgery (late phase). The delayed increase in biopyrrin tended to be higher in patients with UAP, with a simultaneous increase in RNS. The patients in the UAP group had generally high pulmonary capillary wedge pressure (PCWP), although the cardiac index was within a normal range during the delay phase. CONCLUSIONS: The dynamics of biopyrrin levels revealed a biphasic pattern of oxidative stress after OPCAB. Delayed production of oxidative stress may be influenced by preoperative severity of myocardial ischemia and delayed RNS production.

Analysis of oxidative stress expressed by urinary level of biopyrrins and 8-hydroxydeoxyguanosine in patients with chronic schizophrenia.

AIMS: Previous studies have supported the claim that psychological stress induces the production of reactive oxygen species. Several authors have suggested that patients with psychiatric disorders show high levels of oxidative stress markers. We examined different oxidative stress markers in patients with chronic schizophrenia. METHODS: This study included 29 patients with chronic schizophrenia and 30 healthy volunteers. The concentration of urinary biopyrrins and 8-hydroxydeoxyguanosine (8-OHdG), as measured by enzyme-linked immunosorbent assay, were normalized to the urinary concentration of creatinine. Psychiatric symptoms were assessed by the administration of the Brief Psychiatric Rating Scale (BPRS). RESULTS: The concentration of biopyrrins in patients with chronic schizophrenia was significantly higher when compared with healthy volunteers. The correlation between biopyrrin level and the duration of illness was highly significant. There were no significant differences in the levels of urinary 8-OHdG between the two groups. In schizophrenic patients, the level of urinary biopyrrins showed correlations with BPRS scores, while the level of urinary 8-OHdG did not show correlations with BPRS. CONCLUSIONS: Urinary biopyrrins are increased in patients with chronic schizophrenia while urinary 8-OHdG is not increased. These findings suggest that patients with chronic schizophrenia are under the condition of certain oxidative stresses.

Effects of lifestyle factors on urinary oxidative stress and serum antioxidant markers in pregnant Japanese women: A cohort study.

Oxidative stress plays a major pathological role in pregnancy-related complications. Although oxidative stress is induced by exogenous toxins in association with a poor lifestyle in normal subjects, there is little information on the factors altering oxidative stress and antioxidant levels during pregnancy. The purpose of this study was to determine the relationship between lifestyle factors and oxidative stress/antioxidant levels during each trimester and 1-month postpartum. This prospective cohort study followed 54 healthy women through pregnancy; first, second, and third trimester and 1-month postpartum. Participants were administered a questionnaire on characteristics and lifestyle factors. Morning blood and urine samples were obtained to measure urinary biopyrrins and serum coenzyme Q10 (CoQ10) levels. The levels of urinary biopyrrins and serum CoQ10 increased significantly throughout pregnancy, with peak values registered during the third trimester. Higher biopyrrin levels were significantly associated with non-consumption of morning meal during the first trimester, smoking during the third trimester and 1-month postpartum, alcohol consumption during the third trimester, high food-based polyunsaturated fatty acid intake during the third trimester, and poor mental health scores during the first and third trimesters. Higher CoQ10 levels were significantly associated with no smoking during pregnancy and at 1-month postpartum, and with a high frequency of exercise during the third trimester and 1-month postpartum. Thus, pregnancy represents a state of oxidative stress, which can be counterbalanced by increased levels of antioxidants, such as CoQ10. We speculate that certain lifestyle choices such as avoiding smoking can reduce oxidative stress and increase antioxidant levels during pregnancy.

HCV NS3 protease enhances liver fibrosis via binding to and activating TGF-ß type I receptor.

Viruses sometimes mimic host proteins and hijack the host cell machinery. Hepatitis C virus (HCV) causes liver fibrosis, a process largely mediated by the overexpression of transforming growth factor (TGF)-ß and collagen, although the precise underlying mechanism is unknown. Here, we report that HCV non-structural protein 3 (NS3) protease affects the antigenicity and bioactivity of TGF-ß2 in (CAGA)9-Luc CCL64 cells and in human hepatic cell lines via binding to TGF-ß type I receptor (TßRI). Tumor necrosis factor (TNF)-α facilitates this mechanism by increasing the colocalization of TßRI with NS3 protease on the surface of HCV-infected cells. An anti-NS3 antibody against computationally predicted binding sites for TßRI blocked the TGF-ß mimetic activities of NS3 in vitro and attenuated liver fibrosis in HCV-infected chimeric mice. These data suggest that HCV NS3 protease mimics TGF-ß2 and functions, at least in part, via directly binding to and activating TßRI, thereby enhancing liver fibrosis.

Complex of branched cyclodextrin and lidocaine prolonged the duration of peripheral nerve block.

Although laboratories have tried to synthesize new local anesthetics, currently available local anesthetics rarely provide prolonged regional blockade. New models of sustained-release preparations of local anesthetics with liposomes and microspheres have been studied to prolong the duration of the effects of the local anesthetics. In the present study, we examined whether a complex of a branched cyclodextrin (CD), 6-O-alpha-D-maltosyl-beta-cyclodextrin (G2-beta-CD) and lidocaine could prolong local nerve block when compared with plain lidocaine. The sciatic nerve in male Sprague-Dawley rats was blocked with plain lidocaine (n = 10), the complex of G2-beta-CD + lidocaine (n = 10), or plain G2-beta-CD (n = 4). Sensory block was assessed with a hotplate set at 56 degrees C. The median duration of the block was longer in the complex group than in the plain lidocaine group (110 min; range, 70-150 min vs 55 min; range, 40-80 min; P < 0.05), thus demonstrating that the complex with CyD significantly prolonged the nerve block effect of lidocaine. In conclusion, the present study showed that this encapsulating technique with CyD is useful to expand local anesthetic effect in peripheral nerve blockade.

Bilirubin and its oxidative metabolite biopyrrins in patients with acute myocardial infarction.

This study aimed to assess the involvement of bilirubin and its oxidative metabolite biopyrrin in patients with acute myocardial infarction (AMI) and to determine the responsible organs that overproduce these molecules. One hundred thirteen consecutive patients hospitalized for AMI were analyzed. Levels of serum bilirubin, plasma and urinary biopyrrins were measured on the day of admission, day 2, 3, 7 and 14. Expressions of biopyrrins and heme oxygenase-1 (HO-1), a stress-responsive bilirubin-producing enzyme, in heart, aorta, kidney, liver and lung were immunostained with autopsied specimens. Serum bilirubin, plasma and urinary biopyrrins were increased within 24 hr, formed a peak on day 3 and then decreased by day 14. These three parameters were well correlated to each other. The maximum biopyrrin elevation was higher in death cases and associated with impaired left ventricular function. Immunohistochemical analyses revealed biopyrrin accumulation and HO-1 expression in the infracted myocardium. Immunoreactive HO-1 and biopyrrins were also observed in renal tubular cells, aortic wall and lung. Serum bilirubin and its metabolite biopyrrins were elevated in patients with AMI. Plasma and urinary biopyrrin elevation were associated with mortality and morbidity. Induction of anti-oxidative enzyme HO-1 seemed to be involved in the activation of bilirubin/biopyrrin pathway.

Biphasic elevation of bilirubin oxidation during myocardial ischemia reperfusion.

BACKGROUND: The time course of oxidative stress involving nitric oxide (NO) after myocardial ischemia reperfusion (MIR) has not been elucidated in detail, so the present study was designed to assess the dynamics of oxidative stress after MIR, urinary excretion of oxidized bilirubin metabolites (ie, biopyrrins) and their generation in various organs. METHODS AND RESULTS: Rat models of MIR were created by occluding the left coronary artery for 30 min followed by 48 h of reperfusion. Levels of urinary biopyrrins increased biphasically at 8 h and 24 h after MIR. Biopyrrins were upregulated in the lungs at 8 h after MIR, according to immunohistochemistry and ELISA, and at 24 h biopyrrin expression was increased in the heart and lungs. The NO synthase inhibitor, NG-monomethyl-L-arginine, significantly diminished biopyrrin synthesis in the heart and lungs at 24 h, but not in the lungs at 8 h after MIR. Hemodynamic assessment revealed increased left ventricle end-diastolic pressure, suggesting that lung congestion influences pulmonary biopyrrin formation. CONCLUSIONS: The dynamics of urinary biopyrrins might reflect earlier biopyrrin generation in the lungs and delayed formation in both the lungs and heart when NO is involved. Therefore, urinary biopyrrins can serve as a useful marker of systemic oxidative stress after MIR.

5-HT(2C) receptor activation is a common mechanism on proerectile effects of apomorphine, oxytocin and melanotan-II in rats.

5-Hydroxytryptamine (5-HT), dopamine, oxytocin and melanocortin pathways are known to be involved in the induction of penile erections in rats. Although a dopamine-oxytocin-5-HT link in the central nervous system has been suggested to be important to the control of penile erections, the 5-HT receptor subtype that mediates dopamine-oxytocin-5-HT action and the relationship between the dopamine-oxytocin-5-HT pathway and melanocortin pathway have not been fully elucidated. In this study, in order to clarify these matters, we examined the effects of a selective 5-HT(2B)/5-HT(2C) receptors antagonist, 1-(1-methylindol-5-yl)-3-(3-pyridyl)urea (SB200646) and a selective 5-HT(2C) receptor antagonist, 6-chloro-5-methyl-1-[6-(2-methylpyridin-3-yloxy) pyridin-3-yl carbamoyl] indoline (SB242084) on penile erections induced by a dopamine receptor agonist, 10, 11-dihydroxyaporphine (apomorphine), oxytocin, or a melanocortin receptor agonist, melanotan-II (MT-II) in rats. SB200646 at 10 mg/kg and SB242084 at 3 mg/kg, these doses which completely antagonize penile erections induced by 5-HT(2C) receptor agonists, m-chlorophenylpiperazine (mCPP) and (S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine (YM348), significantly inhibited penile erections elicited by apomorphine, oxytocin or MT-II. In addition, in order to clarify further the suggestion that the 5-HT pathway projecting from medulla oblongata to lumbosacral spinal site and lumbosacral 5-HT(2C) receptor are involved in the induction of penile erection, we also examined the proerectile effect of YM348 in spinal and a 5-HT depletor, p-chlorophenyl alanine (pCPA)-treated rats. YM348 induced intracavernous pressure increase in spinal and pCPA-treated rats as well as normal rats. These results suggest that 5-HT(2C) receptor in lumbosacral spinal sites mediates not only dopamine-oxytocin-5-HT action but melanocortin action on penile erections, and that the 5-HT pathway is located downstream from melanocortin pathway as well as the dopamine-oxytocin pathway.

Urinary excretion of oxidative metabolites of bilirubin in subjects with Gilbert syndrome.

BACKGROUND AND AIM: Bilirubin is a potent endogenous antioxidant substance. Recent data suggest a direct relationship exists between urinary excretion of biopyrrins, a novel group of bilirubin oxidative metabolites, and severity of oxidative stress. The aim of this study was to evaluate urinary excretion of biopyrrins in subjects with Gilbert syndrome. METHODS: The study included patients with Gilbert syndrome (n = 33) and healthy blood donors (n = 25). In all subjects complete biochemical tests were conducted along with analysis of urinary excretion of biopyrrins. Linear and logistic regression analyses were used for multiple adjustments of possible confounders/modifiers. RESULTS: As expected, high serum bilirubin levels were found in the Gilbert syndrome group as compared to controls (27.8 +/- 9.7 vs 9.9 +/- 3.0 micromol/L, P < 0.001). In contrast, urinary levels of biopyrrins were substantially lower in the Gilbert syndrome group as compared to normobilirubinemic control subjects (19.9 +/- 26.0 vs 90.2 +/- 139.1 U/g urinary creatinine, P < 0.001). The Gilbert syndrome group also had very low prevalence odds ratios for urinary biopyrrins above the median of the control values even after adjustment for possibly confounding factors (odds ratio 0.18, 95% confidence interval 0.33-0.94; P = 0.042). CONCLUSIONS: An inverse relationship was demonstrated between serum bilirubin level and urinary excretion of biopyrrins, which is presumably due to antioxidative effects of elevated serum bilirubin levels in Gilbert syndrome.

Social stress increases biopyrrins, oxidative metabolites of bilirubin, in mouse urine.

We investigated whether or not three kinds of social stress (isolation, crowding, and confrontation) increase urinary excretion of biopyrrins, oxidative metabolites of bilirubin, in mice. Male BALB/c mice (4 weeks old) were housed 5 per cage for 10 days. After acclimatization, mice were exposed to the above mentioned three kinds of psychosocial stress. Mouse blood and urine samples were collected after 2, 7, and 30 days of stress. Serum levels of corticosterone and urinary levels of biopyrrins were determined by EIA and ELISA, respectively. Adrenal hypertrophy, significant increases in serum concentration of corticosterone and in urinary excretion of biopyrrins were observed in mice exposed to these types of social stress for 7 days. These levels decreased after 30 days, though they were still rather high compared to the control group. These results suggested that social stress causes oxidative stress and that biopyrrins could be useful biomarkers of psychosocial stress.

Characterization of intracavernous pressure increase induced by Ym348, a novel 5-HT2C receptor agonist, in anesthetized rats.

PURPOSE: We investigated the effects of the novel 5-HT2C receptor agonist YM348 [(S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine] on ICP in anesthetized rats and clarified whether behavioral changes such as hypolocomotion induced at a high dose are a cause of the inverted U-shaped PE dose-response curves in conscious rats. MATERIALS AND METHODS: Male Wistar rats (Japan SLC, Shizuoka, Japan) weighing 250 to 315 gm were used. The pro-erectile effect of YM348 (2.03 to 67.7 microg/kg subcutaneously) was examined in conscious rats. ICP was also monitored after YM348 treatment (0.677 to 67.7 mug/kg subcutaneously) in anesthetized rats. Response number, latency, duration, peak pressure and area under the curve were measured. The selective 5-HT2C receptor antagonist SB242084 [(6-chloro-5-methyl-1-[6-(2-methylpyridin-3-yloxy) pyridin-3-yl carbamoyl] indoline)] (0.03 to 1 mg/kg intraperitoneally) was administered 30 minutes before YM348 treatment. RESULTS: YM348 induced PE and ICP increases, and was significantly inhibited by SB242084. Dose-response curves for the number of PEs and ICP increases were an inverted U shape. YM348 decreased the latency of but did not affect the quality of ICP (duration, peak pressure and area under the curve) even at the highest dose. CONCLUSIONS: Activation of 5-HT2C receptor increased ICP and, as a result, induced PE. Since the dose-response curve for the number of ICP increases under anesthetized, behavior independent conditions still showed an inverted U shape, behavioral changes were not likely to have contributed to the inverted U-shaped dose-response curves for the number of PEs in conscious rats. Furthermore, the certain magnitude of ICP increases was likely to have occurred despite the stimulus intensity after the level of 5-HT2C receptor activation crossed the threshold.

Bilirubin exhibits a novel anti-cancer effect on human adenocarcinoma.

Anti-oxidants are essential for intracellular free radical scavenging, as free radicals are one of the causes for tumorigenesis. Our objective was to use bilirubin and investigate its action on human carcinoma cell lines. Bilirubin manifested as a prooxidant showing its cytopathic effect on TMK-1, showing growth inhibition close to 50%. Cell cycle analysis showed an arrest at G0/G1. Flow cytometry investigations with Red CC-1 showed an increase by more than 2 times suggesting a prooxidative role of bilirubin. To check the effect of radicals on DNA, a Comet Assay displayed a typical comet's tail with bilirubin treated slides, only. Further, staining with DAPI showed apoptotic action of bilirubin. Decreased mitochondrial function by bilirubin was observed with Mitotracker Green FM staining. These unexpected data have led us to conclude that bilirubin has anti-cancer activity as a prooxidant and may have a more vital role in the human body than realized.

Carbon monoxide from heme oxygenase-2 Is a tonic regulator against NO-dependent vasodilatation in the adult rat cerebral microcirculation.

Although the brain generates NO and carbon monoxide (CO), it is unknown how these gases and their enzyme systems interact with each other to regulate cerebrovascular function. We examined whether CO produced by heme oxygenase (HO) modulates generation and action of constitutive NO in the rat pial microcirculation. Immunohistochemical analyses indicated that HO-2 occurred in neurons and arachnoid trabecular cells, where NO synthase 1 (NOS1) was detectable, and also in vascular endothelium-expressing NOS3, suggesting colocalization of CO- and NO-generating sites. Intravital microscopy using a closed cranial window preparation revealed that blockade of the HO activity by zinc protoporphyrin IX significantly dilates arterioles. This vasodilatation depended on local NOS activities and was abolished by CO supplementation, suggesting that the gas derived from HO-2 tonically regulates NO-mediated vasodilatory response. Bioimaging of NO by laser-confocal microfluorography of diaminofluorescein indicated detectable amounts of NO at the microvascular wall, the subdural mesothelial cells, and arachnoid trabecular cells, which express NOS in and around the pial microvasculature. On CO inhibition by the HO inhibitor, regional NO formation was augmented in these cells. Such a pattern of accelerated NO formation depended on NOS activities and was again attenuated by the local CO supplementation. Studies using cultured porcine aortic endothelial cells suggested that the inhibitory action of CO on NOS could result from the photo-reversible gas binding to the prosthetic heme. Collectively, CO derived from HO-2 appears to serve as a tonic vasoregulator antagonizing NO-mediated vasodilatation in the rat cerebral microcirculation.

Exclusive association and simultaneous appearance of congophilic plaques and AT8-positive dystrophic neurites in Tg2576 mice suggest a mechanism of senile plaque formation and progression of neuritic dystrophy in Alzheimer's disease.

Progression of neuritic dystrophy is a histological hallmark of Alzheimer's disease (AD) in addition to amyloid deposition and neurofibrillary tangle formation. Dystrophic neurites (DNs) are abnormal neurites, and are closely associated with amyloid deposits. To clarify the process of DN formation, we immunohistochemically investigated phosphorylated tau (AT8 and Ser396)-positive DNs and plaques in Tg2576 mice overexpressing human beta-amyloid precursor protein (APP) with the Swedish type mutation (K670N/M671L). AT8-positive DNs were exclusively associated with the Congo red-positive plaques examined, and all Abeta(1-40)-positive plaques appeared to be associated with AT8-positive DNs, whereas there were no AT8-positive DNs with Abeta(1-42)-positive/Abeta(1-40)-negative plaques. Since we have previously shown that Abeta(1-42)-positive plaque precede Abeta(1-40) deposition, the appearance of congophilic structures is also late. Quantitative analyses were performed on AT8-positive DNs that were associated with congophilic plaques in the cerebral cortex and hippocampus (more than 1,000 plaques). The number of congophilic plaques increased dramatically with age. The area of DNs in the cerebral cortex and hippocampus increased 120- and 60-fold from 11-13 to 20.5 months of age, respectively. Interestingly, the mean ratio of DN area to congophilic plaque area in every plaque was unchanged, approximately 10%, through the ages examined. The mean plaque size was stable with age in both the cortex and hippocampus. These data suggest that the formation of AT8-positive DNs is simultaneous with Congo red-positive plaque development, and that the event may be closely related in the pathological progression of AD.

Effect of YM928, a novel AMPA receptor antagonist, on seizures in EL mice and kainate-induced seizures in rats.

Kainate-induced seizures and seizures induced by tossing stimulation in epilepsy-prone EL mice are considered as models of complex partial seizures. We used these models to evaluate the anticonvulsive effects of 2-[ N-(4-chlorophenyl)- N-methylamino]-4 H-pyrido[3.2-e]-1,3-thiazin-4-one (YM928), a novel alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist. In the kainate-induced seizure test in rats, wet-dog shakes (WDS) were reduced by oral administration of YM928 at doses of 7.5 mg/kg and 30 mg/kg. YM928 (15 mg/kg) reduced the number of WDS within the first 80 min, but then prolonged the time of occurrence compared with the other groups. Significant reduction in kainate-induced motor seizure was observed with 4-30 mg/kg. YM928 did not induce apparently abnormal behaviour at doses of 2-15 mg/kg but did induce sedation at 30 mg/kg. Carbamazepine (40 or 80 mg/kg), valproate (600 mg/kg), diazepam (2.5 mg/kg), and phenobarbital (20 or 40 mg/kg) exerted anticonvulsant effects against motor seizures, but only valproate, at a dose that also caused sedation, suppressed WDS. Phenytoin and ethosuximide did not show significant anti-kainate effects. In the tossing stimulation test in EL mice, i.p. injection of YM928 at 5 mg/kg or 10 mg/kg significantly increased the number of stimulations required to elicit generalized seizure. Carbamazepine (4 or 8 mg/kg), phenytoin (8 or 16 mg/kg), valproate (100-400 mg/kg), diazepam (0.5 mg/kg), phenobarbital (1.3 or 2.5 mg/kg) and ethosuximide (75-300 mg/kg) exerted significant anticonvulsant effects against these seizures. These results indicate that YM928 has anticonvulsant effects on seizure models that are characteristic of partial onset seizures in humans. YM928 is expected to have beneficial effects against human complex partial seizure with secondary generalization or temporal lobe epilepsy.

Suppression of fully kindled seizure and retardation of kindling acquisition by YM928 in the rat kindling model of epilepsy.

We investigated the effects of 2-[N-(4-chlorophenyl)-N-methylamino]-4H-pyrido[3.2-e]-1,3-thiazin-4-one (YM928), a selective alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor antagonist, in the rat kindling model of complex partial seizures. YM928 (10 and 30 mg/kg p.o.) markedly suppressed the motor seizures and afterdischarge induced by electrical stimulation of the amygdala at generalized seizure-triggering threshold intensity. YM928 (10 mg/kg p.o.) did not induce apparent abnormal behavior, but did induce sedation at a dose of 30 mg/kg p.o. YM928 (30 mg/kg p.o.) showed a similar anticonvulsant effect at twice the threshold intensity as it did at threshold intensity. Diazepam (10 mg/kg p.o.) and phenobarbital (60 mg/kg p.o.) also exerted anticonvulsant activities. Diazepam (10 mg/kg) showed a similar effect at twice the threshold as at threshold, but the anticonvulsant effect of phenobarbital (60 mg/kg p.o.) was reversed when the stimulus was doubled. When YM928 (10 mg/kg p.o.) was administered 60 min before daily stimulation of the amygdala, the development of kindling seizure was significantly retarded. These results indicate that YM928 has anticonvulsant effects and suppresses kindling acquisition without sedative effects, and may be suitable as an antiepileptic drug for the treatment of complex partial seizures in humans.

Spinal neurotoxicity and tolerance after repeated intrathecal administration of YM 872, an AMPA receptor antagonist, in rats.

PURPOSE: Although the Alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor antagonist, YM 872, has been considered to be useful in analgesia for both acute and chronic pain, there are no studies of its neurotoxicity and tolerance. We examined the spinal neurotoxicity and tolerance of YM 872 analgesia by repeated intrathecal administration in rats. METHODS: Male Sprague-Dawley rats with lumbar intrathecal catheters received YM 872 at 1 micro g. 10 micro l-1 (eight rats; YM group) or normal saline 10 micro l (eight rats; C group) intrathecally once a day for 30 days. We evaluated the analgesic effects every 3 days, by tail-flick test and behavioral side effects. On the 31st day, the lumbar spinal cord was removed from four randomly selected rats in each group for histological examination. RESULTS: The YM group showed significantly longer tail-flick latency when subjected to a high-intensity light beam than the C group at each measurement time point, although no significant changes in the latency according to the time course of the study were observed for the entire study period of 30 days in either group. No rats showed any side effects. Histologically, only slight lymphocytic cell infiltration and degeneration of myelinated fibers occurred, similarly in both groups. No changes were observed in the spinal cord in either group. CONCLUSION: Administration of YM 872 (1 micro g) once a day for 30 days did not induce any tolerance and caused no histological changes in the spinal cord.

Pharmacological profile of YM348, a novel, potent and orally active 5-HT2C receptor agonist.

YM348, (S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine, showed a high affinity for cloned human 5-HT(2C) receptors (K(i): 0.89 nM). The functional selectivity for 5-HT(2C) receptors in the 5-HT(2) receptor family was the highest among 5-HT(2C) receptor agonists, including m-chlorophenylpiperazine (mCPP) and Ro60-0175 ((S)-2-(6-chloro-5-fluoroindol-1-yl)-1-methylethylamine). Oral administration of YM348 induced penile erections and hypolocomotion in rats, being completely inhibited by a selective 5-HT(2C) receptor antagonist, SB242084 (6-chloro-5-methyl-1-[6-(2-methylpyridin-3-yloxy) pyridin-3-yl carbamoyl] indoline). The dose-response curve for penile erections, unlike that for hypolocomotion, was an inverted U-shape in the dose range of 0.0677-2.03 mg/kg. A selective 5-HT(2A) receptor antagonist, MDL100907 (R(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidine-methanol), and a selective 5-HT(2B) receptor antagonist, RS-127445 (2-amino-4-(4-fluoronaphth-1-yl)-6-isopropylpyrimidine), had no effect on the decline in penile erection frequency at 2.03 mg/kg of YM348. YM348 did not affect blood pressure at 2.03 mg/kg. In conclusion, YM348 is a novel, potent and orally active 5-HT(2C) receptor agonist, and neither the activation of 5-HT(2A) or 5-HT(2B) receptors nor a cardiovascular effect is likely to contribute to the inverted U-shape dose-response curve for penile erections.