European Journal of Pharmacology (v.766, #C)

Inhibition of in vivo [3H]MK-801 binding by NMDA receptor open channel blockers and GluN2B antagonists in rats and mice by Alda Fernandes; Trevor Wojcik; Praveena Baireddy; Rick Pieschl; Amy Newton; Yuan Tian; Yang Hong; Linda Bristow; Yu-Wen Li (1-8).
N-methyl-d-aspartate (NMDA) receptor antagonists, including open channel blockers and GluN2B receptor subtype selective antagonists, have been developed for the treatment of depression. The current study investigated effects of systemically administered NMDA channel blockers and GluN2B receptor antagonists on NMDA receptor activity in rodents using in vivo [3H]MK-801 binding. The receptor occupancy of GluN2B antagonists was measured using ex vivo [3H]Ro 25-6981 binding. Ketamine, a NMDA receptor channel blocker, produced a dose/exposure- and time-dependent inhibition of in vivo [3H]MK-801 binding that was maximal at ~100%. The complete inhibition of in vivo [3H]MK-801 binding was also observed with NMDA receptor channel blockers, AZD6765 (Lanicemine) and MK-801 (Dizocilpine). CP-101,606 (Traxoprodil), a GluN2B antagonist, produced a dose/exposure- and time-dependent inhibition of in vivo [3H]MK-801 binding that was maximal at ~60%. Partial inhibition was also observed with other GluN2B antagonists including MK-0657 (CERC-301), EVT-101, Ro 25-6981 and radiprodil. For all GluN2B antagonists tested, partial [3H]MK-801 binding inhibition was achieved at doses saturating GluN2B receptor occupancy. Combined treatment with ketamine (10 mg/kg, i.p.) and Ro 25-6981(10 mg/kg, i.p.) produced a level of inhibition of in vivo [3H]MK-801 binding that was similar to treatment with either agent alone. In conclusion, this in vivo [3H]MK-801 binding study shows that NMDA receptor activity in the rodent forebrain can be inhibited completely by channel blockers, but only partially (~60%) by GluN2B receptor antagonists. At doses effective in preclinical models of depression, ketamine may preferentially inhibit the same population of NMDA receptors as Ro 25-6981, namely those containing the GluN2B subunit.
Keywords: NMDA; GluN2B; Channel blocker; Antagonist; Radioligand binding; Occupancy;

Ethyl rosmarinate relaxes rat aorta by an endothelium-independent pathway by Piyawadee Wicha; Jiraporn Tocharus; Archawin Nakaew; Rungusa Pantan; Apichart Suksamrarn; Chainarong Tocharus (9-15).
Ethyl rosmarinate is an ester derivative of rosmarinic acid, a major constituent of Hyptis suaveolens. The present study investigated the vasorelaxant mechanism of ethyl rosmarinate in isolated rat aortic rings using an organ bath system. Ethyl rosmarinate (0.1 µM–3 mM) produced concentration-dependent relaxation in aortic rings pre-contracted with phenylephrine (10 µM), exhibiting a pD2 value of 4.56±0.08 and an E max value of 93.82±5.00% (in endothelium-intact rings), as well as a pD2 value of 4.42±0.05 and an E max value of 92.10±3.78% (in endothelium-denuded rings). In the endothelium-denuded rings, the vasorelaxant effect of ethyl rosmarinate was reduced by only 4-aminopyridine (1 mM); however, this was not the case with tetraethylammonium (5 mM), glibenclamide (10 µM), barium chloride (1 mM), and 1H-[1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one (ODQ, 1 µM). Ethyl rosmarinate also reduced the contraction induced by phenylephrine (10 µM) and caffeine (20 mM) in a Ca2+-free solution, and inhibited the contraction induced by increasing extracellular Ca2+ influx, which was induced by KCl (80 mM). Ethyl rosmarinate (10 µM) inhibits concentration-response curves for phenylephrine, while in the same concentration of ethyl rosmarinate has no effect on contractions induced by increasing concentrations of calcium in the presence of high extracellular potassium. Our results suggests that ethyl rosmarinate induces relaxation in aortic rings via an endothelium-independent pathway, which involves the opening of voltage-gated potassium (Kv) channels and the blockade of both Ca2+release from intracellular stores and extracellular Ca2+ influx. Moreover, ethyl-rosmarinate acts on the extracellular Ca2+ influx inhibition by interacting with voltage-operated calcium channels (VOCCs) and receptor-operated calcium channels (ROCCs).
Keywords: Ethyl rosmarinate; Hypertension; Vasorelaxation; Endothelium-independent; Rat aorta;

Obesity is one of the major risk factors for the development of cardiovascular diseases. It is characterized by excessive or abnormal accumulation of adipose tissue, including depots which surround the blood vessels named perivascular adipose tissue (PVAT). PVAT plays endocrine and paracrine roles by producing large numbers of metabolically vasoactive adipokines. The present review outlines our current understanding of the beneficial roles of PVAT in vascular tone and remodeling in healthy subjects supported by clinical studies, highlighting different factors or mechanisms that could mediate protective effects of PVAT on vascular function. Most studies in humans show that adiponectin is the best candidate for the advantageous effect of PVAT. However, in pathological conditions especially obesity-related cardiovascular diseases, the beneficial effects of PVAT on vascular functions are impaired and transform into detrimental roles. This change is defined as PVAT dysfunction. In the current review, the contribution of PVAT dysfunction to obesity-related cardiovascular diseases has been discussed with a focus on possible mechanisms including an imbalance between beneficial and detrimental adipokines (commonly described as decreased levels of adiponectin and increased levels of leptin or tumor necrosis factor-alpha (TNFα)), increased quantity of adipose tissue, inflammation, cell proliferation and endothelial dysfunction. Finally, novel pharmacotherapeutic targets for the treatment of cardiovascular and metabolic disorders are addressed.
Keywords: Perivascular adipose tissue; Vascular tone; Vascular remodeling; Human; Obesity; Atherosclerosis;

The adenosine A2A receptor antagonist, istradefylline improves motor function in patients with advanced Parkinson's disease (PD) optimally treated with a combination of l-DOPA and a dopamine agonist without increasing the risk of troublesome dyskinesia. However, the effects of istradefylline on motor function when administered in combination with low dose of l-DOPA and dopamine agonists as occurs in early PD are unknown. We investigated whether istradefylline enhances the combined anti-parkinsonian effects of a suboptimal dose of l-DOPA and a threshold dose of either the non-ergot dopamine agonist, ropinirole or the ergot dopamine agonist, pergolide in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmoset.Threshold doses of ropinirole (0.025–0.075 mg/kg p.o.) and pergolide (0.01 mg/kg p.o.) produced a weak anti-parkinsonian effect. Co-administration of a suboptimal dose of l-DOPA (2.5 mg/kg p.o.) with threshold doses of the dopamine agonists enhanced their anti-parkinsonian effect that led to increased ‘ON’ time without dyskinesia appearing. Administering istradefylline (10 mg/kg p.o.) with the threshold doses of dopamine agonists and the suboptimal dose of l-DOPA in a triple combination caused a further enhancement of the anti-parkinsonian response but dyskinesia was still absent.In early PD, dopamine agonists are often used as first-line monotherapy, but efficacy is usually lost within a few years, at which time l-DOPA is added but with the risk of dyskinesia appearance. These results show that istradefylline is effective in improving motor function in combination with low dose dopaminergic drug treatment without provoking dyskinesia.
Keywords: Anti-parkinsonian; Adenosine A2A receptor antagonist; L-DOPA; Dopamine agonist; MPTP;

Perindopril and barnidipine alone or combined with simvastatin on hepatic steatosis and inflammatory parameters in hypertensive patients by Giuseppe Derosa; Amedeo Mugellini; Rosa M. Pesce; Angela D’Angelo; Pamela Maffioli (31-36).
The aim of this study was to evaluate the effects of perindopril or barnidipine alone or combined with simvastatin on metabolic parameters and hepatic steatosis degree.One hundred and forty nine mild to moderate hypertensive, normocholesterolemic, overweight or obese outpatients with hepatic steatosis were enrolled. They were treated with perindopril 5 mg/day, or barnidipine, 20 mg/day, for 6 months; subsequently simvastatin, 20 mg/day was added to both treatments for further 6 months. Blood pressure variation was recorded. Patients also underwent an ultrasound examination, at baseline and after 6, and 12 months. We also assessed: fasting plasma glucose (FPG), fasting plasma insulin (FPI), lipid profile, adiponectin (ADN), tumor necrosis factor-α (ΤΝF-α), interleukin-6 (IL-6), high-sensitivity C reactive protein (Hs-CRP).Both perindopril and barnidipine reduced blood pressure, with barnidipine being more effective. Barnidipine, but not perindopril, slightly decreased total cholesterol and triglycerides after 6 months compared to baseline; lipid profile improved in both groups when simvastatin was added. Regarding inflammatory parameters, barnidipine reduced TNF-a, IL-6, and Hs-CRP, both in monotherapy, and after simvastatin addition. Hepatic steatosis parameters improved only when simvastatin was added.We can conclude that barnidipine better reduced blood pressure compared to perindopril and inflammatory parameters. Regarding hepatic steatosis parameters, only the addition of simvastatin improved them.Registration number: NCT02064218, ClinicalTrials.gov Display Omitted
Keywords: Barnidipine; Perindopril; Steatosis;

β-elemene inhibits monocyte–endothelial cells interactions via reactive oxygen species/MAPK/NF-κB signaling pathway in vitro by Meng Liu; Lifei Mao; Abdelkader Daoud; Waseem Hassan; Liangliang Zhou; Jiawei Lin; Jun Liu; Jing Shang (37-45).
The recruitment of monocytes to the active endothelial cells is an early step in the formation of atherosclerotic lesions; therefore, the inhibition of monocyte–endothelial cells interactions may serve as a potential therapeutic strategy for atherosclerosis. Recent studies suggest that β-elemene can protect against atherosclerosis in vivo and vitro; however, the mechanism underlying the anti-atherosclerotic effect by β-elemene is not clear yet. In this study, we aimed to investigate the effects of β-elemene on the monocyte–endothelial cells interactions in the initiation of atherosclerosis in vitro. Our results showed that β-elemene protects human umbilical vein endothelial cells (HUVECs) from hydrogen peroxide-induced endothelial cells injury in vitro. Besides, this molecule inhibits monocyte adhesion and transendothelial migration across inflamed endothelium through the suppression of the nuclear factor-kappa B-dependent expression of cell adhesion molecules. Further, β-elemene decreases generation of reactive oxygen species (ROS) and prevents the activation of mitogen-activated protein kinase (MAPK) signaling pathway in HUVECs. In conclusion, this study would provide a new pharmacological evidence of the significance of β-elemene as a future drug for prevention and treatment of atherosclerosis.
Keywords: β-elemene; Atherosclerosis; Human umbilical vein endothelial cells; Cell adhesion; NF-κB;

Caveolin-1 scaffolding domain residue phenylalanine 92 modulates Akt signaling by Andy E. Trane; Matti A. Hiob; Tanya Uy; Dmitri Pavlov; Pascal Bernatchez (46-55).
Caveolin-1 (Cav-1), the homo-oligomeric coat protein of cholesterol-rich caveolae signalosomes, regulates signaling proteins including endothelial nitric oxide synthase (eNOS). The Cav-1 scaffolding domain (a.a. 82-101) inhibits activated eNOS from producing vascular protective nitric oxide (NO), an enzymatic process involving trafficking and phosphorylation. However, we demonstrated that Cav-1 proteins and peptides bearing F92A substitution (CAVF92A) could promote cardioprotective NO, most likely by preventing inhibition of eNOS by Cav-1. Herein, we showed that wild-type CAV sequence could, similar to CAVF92A, stimulate basal NO release, indicating a need to better characterize the importance of F92 in the regulation of eNOS by Cav-1/CAV. To reduce uptake sequence-associated effects, we conjugated a wild-type CAV derivative (CAVWT) or a F92A variant (CAVF92A) to antennapedia peptide (AP) or lipophilic myristic acid (Myr) and compared their effect on eNOS regulation in endothelial cells. We observed that both CAVWT and CAVF92A could increase basal NO release, although F92A substitution potentiates this response. We show that F92A substitution does not influence peptide uptake, endogenous Cav-1 oligomerization status and Cav-1 and eNOS distribution to cholesterol-enriched subcellular fractions. Instead, F92A substitution in CAVWT influences Akt activation and downstream eNOS phosphorylation status. Furthermore, we show that the cell permeabilization sequence could alter subcellular localization of endogenous proteins, an unexpected way to target different protein signaling cascades. Taken together, this suggests that we have identified the basis for two different pharmacophores to promote NO release; furthermore, there is a need to better characterize the effect of uptake sequences on the cellular trafficking of pharmacophores.
Keywords: Endothelial nitric oxide synthase; Caveolin-1 scaffolding domain; Nitric oxide; Antennapedia peptide; Myristic acid;

Inhibitory effect of fluvoxamine on β-casein expression via a serotonin-independent mechanism in human mammary epithelial cells by Takeshi Chiba; Tomoji Maeda; Soichiro Kimura; Yasunori Morimoto; Atsushi Sanbe; Hideo Ueda; Kenzo Kudo (56-62).
Selective serotonin reuptake inhibitors (SSRIs) are widely used as a first-line therapy in postpartum depression. The objective of this study was to determine the mechanism underlying the inhibitory effects of the SSRI, fluvoxamine, on β-casein expression, an indicator of lactation, in MCF-12A human mammary epithelial cells. Expression levels of serotonin (5-hydroxytryptamine; 5-HT) transporter, an SSRI target protein, and tryptophan hydroxylase 1, a rate-limiting enzyme in 5-HT biosynthesis, were increased in MCF-12A cells by prolactin treatment. Treatment with 1 μM fluvoxamine for 72 h significantly decreased protein levels of β-casein and phosphorylated signal transducer and activator transcription 5 (pSTAT5). Extracellular 5-HT levels were significantly increased after exposure to 1 μM fluvoxamine, in comparison with those of untreated and vehicle-treated cells; however, extracellular 5-HT had little effect on the decrease in β-casein expression. Expression of glucose-related protein 78/binding immunoglobulin protein, a regulator of endoplasmic reticulum (ER) stress, was significantly increased after treatment with 1 μM fluvoxamine for 48 h. Exposure to tunicamycin, an inducer of ER stress, also decreased expression of β-casein and pSTAT5 in a manner similar to fluvoxamine. Our results indicate that fluvoxamine suppresses β-casein expression in MCF-12A cells via inhibition of STAT5 phosphorylation caused by induction of ER stress. Further studies are required to confirm the effect of fluvoxamine on the function of mammary epithelial cells.
Keywords: Selective serotonin reuptake inhibitor; Serotonin; β-casein; Mammary gland; Endoplasmic reticulum stress; MCF-12A;

AG490, a Jak2 inhibitor, suppressed the progression of murine ovarian cancer by Aya Kobayashi; Yuko Tanizaki; Akihiko Kimura; Yuko Ishida; Mizuho Nosaka; Saori Toujima; Yumi Kuninaka; Sawako Minami; Kazuhiko Ino; Toshikazu Kondo (63-75).
Ovarian cancer is the major cause of cancer death among female genital malignancies, and requires developing novel therapeutic measures. Immune escape and acquisition of tolerance by tumor cells are essential for cancer growth and progression. An immunoregulatory enzyme indoleamine 2,3-dioxygenase (IDO) overexpression in tumors is essential for host immune tolerance. Janus-activated kinase-signal transducer and activator of transcription (JAK–STAT) pathway is involved in various kinds of tumor biology. Thus, we examined the effects of STAT1 inhibition by AG490 (a JAK2 inhibitor) on ovarian cancer progression in mice. In vitro study, IFN-γ treatment up-regulated Ido mRNA expression with STAT1 activation in OV2944-HM-1 cells, whereas AG490 treatment significantly inhibited this effect with the suppression of STAT1 phosphorylation. In vivo model, OV2944-HM-1 cells were intraperitoneally/subcutaneously transplanted into syngeneic immunocompetent female mice. AG490 treatment significantly suppressed subcutaneous tumor growth, compared with control. Consistently, in mice intraperitoneally inoculated HM-1 cells, the same treatment significantly improved survival rate with the reduced number of intraperitoneal tumors. Actually, intratumoral IDO expression was significantly suppressed with the reduction of STAT1 activation in AG490-treated mice. Moreover, in tumor microenvironment of mice treated with AG490, the accumulation of anti-tumor leukocytes such as CD8+ T-cells, M1 macrophages, and NK cells was apparently exaggerated with the reciprocal reduction of regulatory T cells. Furthermore, intratumoral expression of anti-tumor cytokines such as IL-1α, IL-1β and IL-12 expression was significantly enhanced in mice treated with AG490. Collectively, JAK/STAT signal pathways may be good molecular target for immunotherapy of ovarian cancer.
Keywords: Ovarian cancer; Peritoneal dissemination; Immunotherapy; JAK/STAT signal; Immune tolerance;

Deficiency in apolipoprotein A-I ablates the pharmacological effects of metformin on plasma glucose homeostasis and hepatic lipid deposition by Eleni A. Karavia; Aikaterini Hatziri; Christina Kalogeropoulou; Nikolaos I Papachristou; Eva Xepapadaki; Caterina Constantinou; Anastasios Natsos; Peristera-Ioanna Petropoulou; Shlomo Sasson; Dionysios J Papachristou; Kyriakos E. Kypreos (76-85).
Recently, we showed that deficiency in apolipoprotein A-I (ApoA-I) sensitizes mice to diet-induced obesity, glucose intolerance and NAFLD. Here we investigated the potential involvement of ApoA-I in the pharmacological effects of metformin on glucose intolerance and NAFLD development. Groups of apoa1-deficient (apoa1 -/- ) and C57BL/6 mice fed western-type diet were either treated with a daily dose of 300 mg/kg metformin for 18 weeks or left untreated for the same period. Then, histological and biochemical analyses were performed. Metformin treatment led to a comparable reduction in plasma insulin levels in both C57BL/6 and apoa1 -/- mice following intraperitoneal glucose tolerance test. However, only metformin-treated C57BL/6 mice maintained sufficient peripheral insulin sensitivity to effectively clear glucose following the challenge, as indicated by a [3H]-2-deoxy-D-glucose uptake assay in isolated soleus muscle. Similarly, deficiency in ApoA-I ablated the effect of metformin on hepatic lipid deposition and NAFLD development. Gene expression analysis indicated that the effects of ApoA-I on metformin treatment may be independent of adenosine monophosphate-activated protein kinase (AMPK) activation and de novo lipogenesis. Interestingly, metformin treatment reduced mitochondrial oxidative phosphorylation function only in apoa1 -/- mice. Our data show that the role of ApoA-I in diabetes extends to the modulation of the pharmacological actions of metformin, a common drug for the treatment of type 2 diabetes.
Keywords: Apolipoprotein A-I; Metformin; HDL lipoprotein; Nonalcoholic fatty liver disease; Obesity; Glucose intolerance;

Gender-related drug effect on several markers of oxidation stress in diabetes patients with and without complications by Maura Lodovici; Elisabetta Bigagli; Cristina Luceri; Edoardo Mannucci; Carlo Maria Rotella; Laura Raimondi (86-90).
We previously reported that circulating lipid (malondialdehyde, MDA) and protein oxidation (carbonyl residues, CO) products can be used as markers of risk for complications in poorly controlled type 2 diabetics. Now, we aimed to evaluate the existence of a gender effect on classical disease markers and oxidative stress parameters and on the effectiveness of metformin and/or statins in reducing CV risk in poorly controlled type 2 diabetics with and without complications. Our results show that diabetics with complications had higher plasma levels of FRAP, SOD and hs-CRP than those without complications, with FRAP and SOD found increased in both genders. Interestingly, male and female patients with complications had higher plasma levels of hs-CRP and MDA respectively, over patients without complications. Multivariate analysis indicated metformin and statin treatments effective in reducing plasma hs-CRP only in female and not in male diabetics with complications. In these latter females, a positive correlation between hs-CRP and triglycerides (TG) levels was found suggesting a causal relationship between them. Statin treatment was effective in reducing MDA in diabetics with complications irrespective of the gender. These data support the addition of statins to diabetic standard therapy to control oxidation injury and inflammation and, for the first time, indicate female patients with complications more responsive than males to the CV protection offered by metformin.
Keywords: Type 2 diabetes; Oxidation stress; Hs-CRP; Gender; Metformin; Statins;

Beneficial effect of magnesium lithospermate B on cerebral ischemia–reperfusion injury in rats involves the regulation of miR-107/glutamate transporter 1 pathway by Zhong-Bao Yang; Xiu-Ju Luo; Kai-Di Ren; Jing-Jie Peng; Bin Tan; Bin Liu; Zheng Lou; Xiao-Ming Xiong; Xiao-Jie Zhang; Xian Ren; Jun Peng (91-98).
Recent studies uncovered that glutamate accumulation following cerebral ischemia–reperfusion (I/R) was related to the dysfunction of miR-107/glutamate transporter-1(GLT-1) pathway and magnesium lithospermate B (MLB) possesses the pharmacological activity of anti-excitotoxicity. This study aims to explore whether MLB is able to protect rat brain from excitatory neurotoxicity during I/R by modulating miR-107/GLT-1 pathway. Rats were subjected to 2 h of cerebral ischemia following by 24 h of reperfusion to establish an I/R injury model, which showed an increase in neurological deficit score, infarct volume and cellular apoptosis concomitant with glutamate accumulation, miR-107 elevation and GLT-1 down-regulation. Administration of MLB reduced I/R-induced cerebral injury accompanied by a reverse in glutamate accumulation, miR-107 and GLT-1 expression. Next, we examined the association of MLB with miR-107/GLT-1 pathway in a nerve cell hypoxia/reoxygenation (H/R) injury model. H/R treatment increased the nerve cells apoptosis concomitant with glutamate accumulation and miR-107 elevation, and suppressed GLT-1 expression, mimicking our in vivo findings. All these effects were reversed in the presence of MLB, confirming a strong correlation between MLB and miR-107/GLT-1 pathway. Based on these observations, we conclude that MLB is able to protect the rat brain from excitatory neurotoxicity during I/R through the regulation of miR-107/GLT-1 pathway.
Keywords: Magnesium lithospermate B; miR-107; Glutamate transporter 1; Brain; Ischemia–reperfusion;

The effects of caffeic, coumaric and ferulic acids on proliferation, superoxide production, adhesion and migration of human tumor cells in vitro by Nouha Nasr Bouzaiene; Soumaya Kilani Jaziri; Hervé Kovacic; Leila Chekir-Ghedira; Kamel Ghedira; José Luis (99-105).
Reactive oxygen species are well-known mediators of various biological responses. In this study, we examined the effect of three phenolic acids, caffeic, coumaric and ferulic acids, on superoxide anion production, adhesion and migration of human lung (A549) and colon adenocarcinoma (HT29-D4) cancer cell lines. Proliferation of both tumor cells was inhibited by phenolic acids. Caffeic, coumaric and ferulic acids also significantly inhibited superoxide production in A549 and HT29-D4 cells. Superoxide anion production decreased by 92% and 77% at the highest tested concentration (200 µM) of caffeic acid in A549 and HT29-D4 cell lines respectively. Furthermore, A549 and HT29-D4 cell adhesion was reduced by 77.9% and 79.8% respectively at the higher tested concentration of ferulic acid (200 µM). Migration assay performed towards A549 cell line, revealed that tested compounds reduced significantly cell migration. At the highest concentration tested (200 µM), the covered surface was 7.7%, 9.5% and 35% for caffeic, coumaric or ferulic acids, respectively. These results demonstrate that caffeic, coumaric and ferulic acids may participate as active ingredients in anticancer agents against lung and colon cancer development, at adhesion and migration steps of tumor progression.
Keywords: Reactive oxygen species; Phenolic acids; Adhesion; Migration;

Schizophrenia is a complex and chronic mental health disease that affects nearly 1% of the population worldwide. While the current antipsychotic medications have profoundly impacted the treatment of schizophrenia over the past 50 years, the newer atypical antipsychotics have not fulfilled initial expectations, and enormous challenges remain in long-term treatment of this debilitating disease. In particular, improved treatment of the negative symptoms and cognitive dysfunction in schizophrenia which greatly impact overall morbidity is required. Nitric oxide (NO) is considered as an intra- and inter-cellular messenger in the brain. The implication of NO in the pathogenesis of schizophrenia is documented. Specifically, underproduction of NO is linked to this pathology. This, in turn, indicates that enhancement of nitrergic activity might be beneficial in this disease. Therefore, novel molecules aiming to increase NO production such as NO donors might constitute potential candidates for the treatment of schizophrenia. Here I intended to critically review advances in research of these emerging molecules for the treatment of this psychiatric disorder. Present analysis suggests that NO donors might be a promising class of compounds for the treatment of schizophrenia. However, the potential neurotoxicity and the narrow therapeutic window of NO donors would add a note of caution in this context.
Keywords: Nitric oxide; Nitric oxide donor; Schizophrenia;

Chronic inflammation, imbalance between the extracellular matrix synthesis and degradation, and loss of vascular smooth muscle cells (SMCs) contribute to the development of abdominal aortic aneurysm (AAA). The purpose of this study was to investigate the effect of the therapy with periaortic incubation of c-Jun N-terminal protein kinase inhibitor SP600125 infused from an osmotic pump and subadventitial injection of lysyl oxidase (LOX) gene modified autologous smooth muscle progenitor cells (SPCs) on treatment of AAA in a rabbit model. Obvious dilation of the abdominal aorta in the control group was caused by periaortic incubation of calcium chloride and elastase. But the progression of aortic dilation was significantly decreased after the treatment with SP600125 and LOX gene modified SPCs compared to the treatment with phosphate-buffered saline. This therapy could inhibit matrix metalloproteinases expression, enhance elastin synthesis, improve preservation of elastic laminar integrity, benefit SPCs survival and restore SMCs population. It seemed that this method might provide a novel therapeutic strategy to treat AAA.
Keywords: Abdominal aortic aneurysm; C-Jun N-terminal protein kinase; Drug delivery system; Smooth muscle progenitor cell; SP600125;

Bidirectional effects of dexmedetomidine on human platelet functions in vitro by Shuji Kawamoto; Hideo Hirakata; Naoko Sugita; Kazuhiko Fukuda (122-128).
Platelets express the imidazoline (I)-receptor, I1 and I2, as well as the α2-adrenoceptor. Although dexmedetomidine, a selective α2-adrenoceptor agonist with some affinity for the I-receptor is expected to affect platelet function, the effects of dexmedetomidine on platelet functions remain unclear. In the present study, we investigated the effects of dexmedetomidine on human platelet functions in vitro. The effects of dexmedetomidine on platelet aggregation were examined using aggregometers. The formation of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) in platelets was measured by an enzyme immunoassay. In addition, P-selectin expression in platelets was estimated by flow cytometry. We showed that dexmedetomidine enhances platelet aggregation. But in the presence of yohimbine, an α2-antagonist, dexmedetomidine suppressed platelet aggregation. Efaroxan, an I1-antagonist, and methylene blue, a soluble guanylate cyclase inhibitor, abolished the suppressive effect of dexmedetomidine, whereas idazoxan, an I2-antagonist, showed no effect. Dexmedetomidine suppressed cAMP formation and enhanced P-selectin expression in platelets, and these effects were inhibited by yohimbine. Dexmedetomidine increased cGMP formation in platelets in the presence of yohimbine, and this increase was suppressed by efaroxan. These results demonstrated that dexmedetomidine has both enhancing and suppressive effects on human platelet functions through its action on the α2-adrenoceptor and on the I1-imidazoline receptor, respectively.
Keywords: Dexmedetomidine; Platelet; α2-Adrenoceptor; Imidazoline receptor;

In the present study, we evaluated the in vitro characteristics of bixalomer for phosphate binding and swelling and assessed the urinary phosphorus excretion and plasma phosphorus level-lowering effect of bixalomer. The maximum phosphate binding capacity was 6.49 mmol/g and was maximized at pH 6.09. In rats, consuming a high-phosphorus diet resulted in elevated urinary phosphorus excretion, while consuming a diet of bixalomer (0.3–9%) or sevelamer hydrochloride (sevelamer HCl; 3–9%) mixed with a high-phosphorus diet resulted in a dose-dependent reduction in urinary phosphorus excretion. Rats with adenine sulfate-induced chronic kidney disease (CKD) had plasma phosphorus levels of 14.9–18.8 mg/dl, while CKD rats administered a 3% bixalomer or 3% sevelamer HCl diet for 4 weeks had relatively decreased plasma phosphorus levels (6.86±1.42 or 5.32±0.27 mg/dl, respectively). Bixalomer elevated the lowered blood pH in acidemic CKD rats, while sevelamer HCl administration only exacerbated the acidemia. The swelling index, which represents water adsorption capacity, of bixalomer was measured by subtracting the dry weight from the hydrated wet weight of the polymer. The swelling index of bixalomer was four times lower than that of sevelamer HCl. Bixalomer was found to reduce the plasma phosphorus level in CKD rats by binding phosphate in the small intestine and reducing phosphate absorption. Bixalomer showed favorable characteristics of a smaller swelling index than sevelamer HCl and amelioration of metabolic acidosis. These findings suggest that bixalomer may be useful in treating hyperphosphatemia, with fewer gastrointestinal side effects and amelioration of metabolic acidosis than sevelamer HCl.
Keywords: Hyperphosphatemia; Phosphate binder; Bixalomer; Swelling index; Metabolic acidosis;

Evaluation of the abuse potential of AM281, a new synthetic cannabinoid CB1 receptor antagonist by Chrislean Jun Botanas; June Bryan de la Peña; Irene Joy dela Pena; Reinholdgher Tampus; Hee Jin Kim; Seong Shoon Yoon; Joung-Wook Seo; Eun Ju Jeong; Jae Hoon Cheong (135-141).
AM281 (1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-4-morpholinyl-1H-pyrazole-3-carboxamide) is a new synthetic cannabinoid CB1 receptor antagonist. Similar to other cannabinoid antagonists, AM281 has been suggested to have therapeutic indications. However, recent reports have suggested that cannabinoid CB1 receptor antagonists may share similar behavioral effects with other drugs of abuse such as cocaine and amphetamine. These reports cast doubts on the safety profile of AM281. Thus, in the present study we evaluated the abuse potential (rewarding and reinforcing effects) of AM281 through two of the most widely used animal models for assessing the abuse potential of drugs: the conditioned place preference (CPP) and self-administration (SA) tests. Experiments were performed in Sprague–Dawley rats in various dosages [CPP (0.1, 0.5 or 2.5 mg/kg), SA (0.005, 0.025 or 0.1 mg/kg/infusion)]. We also delved into the consequences of repeated drug exposure on the subsequent response to the drug. Thus, parallel experiments were carried out in rats pretreated with AM281 for 7 or 14 days. Our findings indicated that AM281, at any dose, did not induce CPP and SA in drug-naïve rats. Interestingly, significant CPP (0.5 mg/kg of AM281), but not SA, was observed in 14 days pretreated rats. These observations suggest that AM281 per se has no or minimal rewarding and reinforcing properties, but alterations in neuronal functions and behavior due to repeated AM281 exposure may contribute in part to the abuse potential of this drug. In view of this finding, we advocate the careful use, monitoring, and dispensation of AM281.
Keywords: AM281; Cannabinoid antagonist; Abuse potential; Conditioned place preference; Self-administration;

Expression of heat shock protein 27 and troponin T and troponin I after naloxone-precipitated morphine withdrawal by Elena Martínez-Laorden; Pilar Almela; María-Victoria Milanés; María-Luisa Laorden (142-150).
Heat shock protein (Hsp27) renders cardioprotection from stress situations but little is known about its role in myofilaments. In this study we have evaluated the relationship between Hsp27 and troponin response after naloxone-induced morphine withdrawal. Rats were treated with two morphine (75 mg) pellets during six days. Precipitated withdrawal was induced by naloxone on day seven. Hsp27 expression, Hsp27 phosphorylated at serine 82 (Ser82), cardiac troponin T (cTnT), cardiac troponin I (cTnI) and µ-calpain were evaluated by immunoblotting in left ventricle. Hsp, cTnT and cTnI was also evaluated by immunofluorescence procedure. Our results show that enhancement in Hsp27 expression and phosphorylation induced by naloxone-precipitated morphine withdrawal occurs with concomitant increases of cTnT and µ-calpain expression, whereas cTnI was decreased. We also observed co-localization of Hsp27 with cTnT in cardiac tissues. These findings provide new information into the possible role of Hsp27 in the protection of cTnT degradation by µ-calpain (a protease mediating proteolysis of cTnT and cTnI) after morphine withdrawal.
Keywords: Hsp 27; µ-calpain; left ventricle; Morphine withdrawal; Troponins.;

Nociceptin/orphanin FQ receptor and pain: Feasibility of the fourth opioid family member by Lorenzo Di Cesare Mannelli; Laura Micheli; Carla Ghelardini (151-154).
The pharmacological management of chronic pain is a major therapeutic problem. The need of repeated treatments reduces the usefulness of classical analgesic drugs, like μ opioid receptor (MOP) agonists, characterized by tolerance development, side effects and abuse. Moreover, the pathological persistence of pain modifies nociceptive signals and pain-devoted structure activity weakening MOP agonists and making difficult the research of new active molecules. Nociceptine/orphanin FQ (N/OFQ) and its receptor (NOP) offers a peculiar opioid system able to interact with MOP receptors and made more sensitive by chronic pain conditions. The pain reliever efficacy of NOP agonists against persistent pain, mainly neuropathic pain, has been highlighted after intrathecal infusions in rats and non human primates (NHPs). The differences emerged between the effects of NOP stimulation in rodents and NHPs allow to hypothesize the relevance of NOP modulators in higher organisms strongly encouraging the development of compounds active by a systemic route. Possible applicative perspectives are (i) selective NOP agonists as such, (ii) NOP modulation as adjuvant of MOP-based treatments, or (iii) mixed non-selective agonists vs NOP and classical opioid receptors.
Keywords: NOP; MOP; Morphine; Chronic pain; Neuropathic pain;

Intrathecal administration of nociceptin/orphanin FQ receptor agonists in rats: A strategy to relieve chemotherapy-induced neuropathic hypersensitivity by Laura Micheli; Lorenzo Di Cesare Mannelli; Anna Rizzi; Remo Guerrini; Claudio Trapella; Girolamo Calò; Carla Ghelardini (155-162).
Oxaliplatin and paclitaxel are considered central components in the treatment of colorectal and breast cancer, respectively. The development of neuropathy during chronic treatment represents the major dose-limiting side effect that leads to discontinuation or interruption of therapies. The management of neuropathy is a challenge to individuate innovative therapeutic strategies based on new targets and correct routes of administration. We evaluated the hypersensitivity reliever effect of different opioid receptor agonists in rat models of oxaliplatin and paclitaxel-induced neuropathy. Compounds were spinally infused by intrathecal catheter. In oxaliplatin-treated rats, 0.3 nmol morphine induced the reversion of the mechanical hypersensitivity (Paw-pressure test), nociceptin/orphanin FQ (N/OFQ; 0.3–3 nmol) significantly increased the pain threshold without reaching the values of the control animals. The N/OFQ peptide (NOP) receptor full agonist UFP-112 reverted pain threshold alterations at lower dosage (0.1 nmol) vs morphine and N/OFQ, the partial agonist UFP-113 (0.1–1 nmol) was similar to N/OFQ. The higher efficacy of morphine vs N/OFQ was highlighted also in paclitaxel-treated rats. The mechanical hypersensitivity was fully reverted by 0.1 nmol UFP-112 and UFP-113. In conclusion, intrathecal μ opioid peptide (MOP) and NOP receptor agonists relieved chemotherapy-induced neuropathic pain. The synthetic peptides showed valuable potency and efficacy suggesting the NOP system as an exploitable target.
Keywords: Chemotherapy-induced neuropathic pain; Intrathecal catheterization; NOP agonists; Oxaliplatin; Paclitaxel; Opioids;