Methylglyoxal Enhances Sodium Nitroprusside–Induced Relaxation in Rat Aorta

The concentration of methylglyoxal (MGO), a metabolite of glucose, increases in plasma of type II diabetic patients as well as in tissues of hypertensive rats. We have previously shown that MGO inhibited noradrenaline (NA)–induced smooth muscle contraction in rat aorta. However, the effect of MGO on relaxing responses in isolated blood vessel remains to be clarified. Thus, we examined if MGO affects acetylcholine (ACh)– or sodium nitroprusside (SNP)– induced vasodilation on NA (100 nM)–induced pre-contraction in rat thoracic aorta. Treatment of endothelium-intact aorta with MGO (420 μM, 30 min) did not change ACh (1 nM – 3 μM)–induced endothelium-dependent relaxation. In contrast, treatment of endothelium-denuded aorta with MGO shifted the concentration–response curve for SNP (0.1 – 300 nM) to the left. MGO increased reactive oxygen species (ROS) production in smooth muscle on analysis of protein carbonylation. Anti-oxidant agents such as tempol (10 μM), catalase (5000 U/mL), and nitric oxide synthase inhibitor, N^G-nitro-l-arginine methylester (100 μM) had no effect on MGO-induced enhancement of SNP-induced relaxation. However, iberiotoxin (100 nM), a large-conductance Ca²⁺-activated K⁺ (BK _Ca)–channel inhibitor, significantly prevented the effect. The present study revealed that MGO enhanced SNP-induced relaxation in a ROS-independent manner via in part opening smooth muscle BK _Ca channels.     

Butea superba–Induced Amelioration of Cognitive and Emotional Deficits in Olfactory Bulbectomized Mice and Putative Mechanisms Underlying Its Actions

This study investigated the effects of alcoholic extract of Butea superba (BS) on cognitive deficits and depression-related behavior using olfactory bulbectomized (OBX) mice and the underlying molecular mechanisms of its actions. OBX mice were treated daily with BS (100 and 300 mg/kg, p.o.) or reference drugs, tacrine (2.5 mg/kg, i.p.) and imipramine (10 mg/kg, i.p.) from day 3 after OBX. OBX impaired non-spatial and spatial cognitive performances, which were elucidated by the novel object recognition test and modified Y maze test, respectively. These deficits were attenuated by tacrine and BS but not imipramine. OBX animals exhibited depression-like behavior in the tail suspension test in a manner reversible by imipramine and BS but not tacrine. OBX down-regulated phosphorylation of synaptic plasticity–related signaling proteins: NMDA receptor, AMPA receptor, calmodulin-dependent kinase II, and cyclic AMP-responsive element-binding protein. OBX also reduced choline acetyltransferase in the hippocampus. BS and tacrine reversed these neurochemical alterations. Moreover, BS inhibited ex vivo activity of acetylcholinesterase in the brain. These results indicate that BS ameliorates not only cognition dysfunction via normalizing synaptic plasticity–related signaling and facilitating central cholinergic systems but also depression-like behavior via a mechanism differing from that implicated in BS amelioration of cognitive function in OBX animals.     

Mechanisms of Early Afterdepolarlzations and Triggered Activity in Guinea Pig Papillary Muscles Induced by Histamine

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Comparative Evaluation of Calcium-Sensitizing Agents,Pimobendan and SCH00013, on the Myocardial Function of Canine Pacing–Induced Model of Heart Failure

Pimobendan and SCH00013 are calcium sensitizers that possess dual action of calcium sensitization and phosphodiesterase-III inhibition. This study was conducted to comparatively evaluate the effect of these medications on the myocardial function of the canine pacing-induced heart failure model using echocardiography. Heart failure was induced in 20 dogs, to which pimobendan and two different doses of SCH00013 were administered orally to 15 dogs for 3 weeks, and the remaining 5 dogs served as the control. Cardiac evaluations were performed at baseline, week 1, week 2, and week 3. Significant thinning and dilation of the left ventricles, with systolic dysfunction, indicated by reduction of fractional shortening (FS) and strain values, were observed with a low dose of SCH00013. Whereas, although systolic dysfunction was observed with reduction of FS and radial strain, significant dilation and thinning of the left ventricles and reduction of circumferential strain were not observed with pimobendan. Pimobendan had a potent positive inotropic effect, with little effect on synchronicity, while low-dose SCH00013 had a weaker positive inotropic effect but was able to sustain synchronicity. Although, it failed to show significant statistical differences, the results of this study allow speculations that administration of pimobendan and SCH00013 may have differing effect on the myocardial function in the canine pacing-induced heart failure model.     

Affective Disorders and Nitric Oxide: A Role in Pathways to Relapse and Refractoriness?

Glutamatergic mechanisms are powerful activators of thel -arginine-nitric oxide (NO) pathway in the central nervous system (CNS). While these mechanisms have been implicated in a variety of neurodegenerative disorders, as well as psychiatric disorders such as schizophrenia and anxiety, a possible role in affective disorders has not been defined. Low gamma-hydroxy butyric acid (GABA)/high glutamate ratios appear to be aetiological factors in the syndrome of depression. In addition to effects on biogenic amines, typical antidepressants exhibit both glutamate-modulating actions and GABA enhancing properties, whileN-methyl-d -aspartate (NMDA)-receptor antagonists, similarly, display antidepressant efficacy. Excessive activation of glutamatergic/nitrergic mechanisms, leading to limbic and subcortical kindling and the synthesis of specific immediate early genes (IEG) and retrograde messengers such as NO, allow the formation of memory traces which can either predict remission of dysphoric mood or, alternatively, the development of relapse and treatment refractoriness. In parallel to its ability to induce refractoriness, lithium can augment glutamate responses, is proconvulsant, regulates gene expression and has distinct effects on NO and cGMP. These effects, including refractoriness, may be overcome with anticonvulsant/GABA-agonists or an NMDA antagonist. Not only are anti-glutamatergic or GABA-enhancing mechanisms thus vital for successful remission of depression, they may also regulate homeostatic mechanisms preventing dysphoric mood recurrence. The diverse effects of NO and cGMP on subcellular events, the unique physico-chemical properties of NO and its involvement in cellular memory processes and synaptic plasticity makes it an ideal regulator of short- and long-term adaptive changes associated with mood regulation.     

Synapse Loss Induced by Interleukin-1β Requires Pre- and Post-synaptic Mechanisms

Interleukin-1β (IL-1β) is an inflammatory cytokine that exerts marked effects on neuronal function and survival. Here we examined the effects of IL-1β on synapses between rat hippocampal neurons in culture using an imaging-based assay to quantify clusters of the scaffolding protein postsynaptic density 95 fused to green fluorescent protein. Treatment with IL-1β for 24?h induced a 23?±?3% loss in the number of synaptic sites. Pharmacological studies indicated that synapse loss was mediated by the IL-1 receptor with subsequent activation of two pathways. COX2-mediated prostaglandin production and postsynaptic activation of a Src family tyrosine kinase were required. Presynaptic release of glutamate with subsequent activation of NMDA receptors was necessary for IL-1β-induced synapse loss. Neither Src activation nor prostaglandin E2 (PGE2) application alone was sufficient to reduce the number of synapses. However, in cells expressing constitutively active or pharmacologically activated Src, PGE2 induced synapse loss. Thus, IL-1β reduces the number of synaptic connections by simultaneously activating multiple pathways that require both pre- and post-synaptic activity. These results highlight targets that may prove important for pharmacotherapy of neuroinflammatory disease.     

Mechanisms of Chronic Nicotine Treatment–Induced Enhancement of the Sensitivity of Cortical Neurons to the Neuroprotective Effect of Donepezil in Cortical Neurons

We have previously shown that chronic donepezil treatment induces nicotinic acetylcholine receptor up-regulation and enhances the sensitivity of the neurons to the neuroprotective effect of donepezil. Further analyses revealed that the nicotinic receptor is involved in this enhancement. In this study, we examined whether nicotinic receptor stimulation is sufficient to make neurons more sensitive to donepezil. We treated primary cultures of rat cortical neurons with nicotine and confirmed that chronic nicotine treatment induced nicotinic receptor up-regulation and made the neurons more sensitive to the neuroprotective effects of donepezil. Analyses with receptor antagonists and kinase inhibitors revealed that the effects of chronic nicotine treatment are mediated by nicotinic receptors and their downstream effectors including phosphatidylinositol 3-kinase. In contrast to chronic donepezil treatment that enhanced the level of nicotine-induced Ca²⁺ influx, chronic nicotine treatment did not significantly alter the level of Ca²⁺ influx.     

Angiostatin Inhibition of Vascular Endothelial Growth Factor– Stimulated Nitric Oxide Production in Endothelial Cells

Angiostatin (AS), a proteolytic fragment of plasminogen, is a potent antiangiogenic factor. It was reported that AS attenuates the vasodilatory response to vascular endothelial growth factor (VEGF) in isolated interventricular arterioles. Here, we investigated the effect of AS on nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs). AS inhibited VEGF-stimulated NO production in a dose-dependent manner, whereas AS alone did not affect basal NO production. Disruption of kringle structures by reduction of disulfide bonds resulted in the loss of the inhibitory effect of AS on VEGF-stimulated NO production. To elucidate how AS might impair VEGF activation of endothelial NO synthase (eNOS), we further examined whether AS would affect Ca²⁺-dependent and -independent pathways of eNOS activation. AS had no effect on the transient increase in cytosolic Ca²⁺ levels elicited by VEGF. In contrast, AS prevented VEGF-potentiated eNOS phosphorylation at Ser1177. These results clearly indicate that AS inhibits VEGF-stimulated NO production in HUVECs without affecting basal NO production. The kringle structures of AS are required for this effect, and impairment of Ser1177 phosphorylation of eNOS might be involved in the inhibition of VEGF-stimulated NO production by AS.     

Effects of Aspirin and Mefenamic Acid on Soman Poisoning–Induced Neuropathology in Mice

The efficacy of aspirin and mefenamic acid to counteract soman-induced brain damage was investigated in mice. Neuronal damage was evaluated in the hippocampus and amygdala by performing ω3 receptor density measurements and hemalun–phloxin staining. The effect of both drugs on the proliferation of neural progenitors after soman exposure was also assessed. Mefenamic acid aggravated the soman-induced hippocampal neuropathology. On the other hand, aspirin recorded a weak neuroprotective effect in the amygdala. However, this drug also diminished the proliferation of neural precursor cells. The possible neurochemical mechanisms underlying such differences in the efficacy of the two drugs are also reviewed.     

Multiple Inhibition Mechanisms and Prediction of Drug–Drug Interactions: Status of Metabolism and Transporter Models as Exemplified by Gemfibrozil–Drug Interactions

Purpose To assess the consequences of multiple inhibitors and differential inhibition mechanisms on the prediction of 12 gemfibrozil drug–drug interactions (DDIs). In addition, qualitative zoning of transporter-related gemfibrozil and cyclosporine DDIs was investigated. Methods The effect of gemfibrozil and its acyl-glucuronide on different enzymes was incorporated into a metabolic prediction model. The impact of CYP2C8 time-dependent inhibition by gemfibrozil acyl-glucuronide was assessed using repaglinide, cerivastatin, loperamide, rosiglitazone and pioglitazone DDIs. Gemfibrozil and cyclosporine inhibition data obtained in human embryonic kidney cells expressing OATP1B1 and hepatic input concentration ([I]in) were used for qualitative zoning of 14 transporter-mediated DDIs. Results Incorporation of time-dependent inhibition by gemfibrozil glucuronide showed no significant improvement in the prediction, as CYP2C8 contributed <65% to the overall elimination of the victim drugs investigated. Qualitative zoning of OATP1B1 DDIs resulted in no false negative predictions; yet the magnitude of observed interactions was significantly over-predicted. Conclusions Time-dependent inhibition by gemfibrozil glucuronide is only important for victim drugs eliminated predominantly (>80%) via CYP2C8. Qualitative zoning of OATP1B1 inhibitors based on [I]in/K _i is valid in drug screening to avoid false negatives. Refinement of the transporter model by incorporating the fraction of drug transported by a particular transporter is recommended.     

Mechanisms and pharmacology of diabetic neuropathy – experimental and clinical studies

Neuropathic pain is the most common chronic complication of diabetes mellitus. The mechanisms involved in the development of diabetic neuropathy include changes in the blood vessels that supply the peripheral nerves; metabolic disorders, such as the enhanced activation of the polyol pathway; myo-inositol depletion; and increased non-enzymatic glycation. Currently, much attention is focused on the changes in the interactions between the nervous system and the immune system that occur in parallel with glial cell activation; these interactions may also be responsible for the development of neuropathic pain accompanying diabetes. Animal models of diabetic peripheral neuropathy have been utilized to better understand the phenomenon of neuropathic pain in individuals with diabetes and to define therapeutic goals. The studies on the effects of antidepressants on diabetic neuropathic pain in streptozotocin (STZ)-induced type 1 diabetes have been conducted. In experimental models of diabetic neuropathy, the most effective antidepressants are tricyclic antidepressants, selective serotonin reuptake inhibitors, and serotonin norepinephrine reuptake inhibitors. Clinical studies of diabetic neuropathy indicate that the first line treatment should be tricyclic antidepressants, which are followed by anticonvulsants and then opioids. In this review, we will discuss the mechanisms of the development of diabetic neuropathy and the most common drugs used in experimental and clinical studies.     

Molecular and biopharmaceutical investigation of alginate–inulin synbiotic coencapsulation of probiotic to target the colon

Colon targeting, as a site-specific delivery for oral formulation, remains a major challenge, especially for sensitive bioactive components such as therapeutic forms of phages, live attenuated virus and prebiotics–probiotics association. Synbiotics could be used to protect encapsulated probiotics during the gastrointestinal tract and control their release in the colon. To achieve these goals, effective prebiotics, such as inulin, could be combined with alginate – the most exploited polymer used for probiotic encapsulation – in the form of beads. This work aimed to study the biopharmaceutical behaviour of alginate beads (A) and inulin–alginate beads of different inulin concentrations (5 or 20%) in 2% alginate (AI5, AI20). Beads were loaded with three probiotic strains (Pediococcus acidilactici Ul5, Lactobacillus reuteri and Lactobacillus salivarius). Dissolution of beads was studied by USP4 under conditions simulating the gastrointestinal condition. The survival rates of the bacterial strains were measured by a specific qPCR bacterial count. Mucoadhesiveness of beads was studied by an ex vivo method using intestinal mucosa. To understand the behaviour of each formulation, the ultrastructure of the polymeric network was studied using scanning electron microscopy (SEM). Molecular interactions between alginate and inulin were studied by Fourier transform infra-red spectroscopy (FTIR). Dissolution results suggested that the presence of inulin in beads provided more protection for the tested bacterial strains against the acidic pH. AI5 was the most effective formulation to deliver probiotics to the colon simulation conditions. FTIR and SEM investigations explained the differences in behaviour of each formula. The developed symbiotic form provided a promising matrix for the development of colonic controlled release systems.