Clinical, virological and epidemiological characterization of dengue outbreak in Myanmar, 2015.

Hospital-based surveillance was conducted at two widely separated regions in Myanmar during the 2015 dengue epidemic. Acute phase serum samples were collected from 332 clinically diagnosed dengue patients during the peak season of dengue cases. Viremia levels were measured by quantitative real-time PCR and plaque assays using FcγRIIA-expressing and non-FcγRIIA-expressing BHK cells to specifically determine the infectious virus particles. By serology and molecular techniques, 280/332 (84·3%) were confirmed as dengue patients. All four serotypes of dengue virus (DENV) were isolated from among 104 laboratory-confirmed patients including two cases infected with two DENV serotypes. High percentage of primary infection was noted among the severe dengue patients. Patients with primary infection or DENV IgM negative demonstrated significantly higher viral loads but there was no significant difference among the severity groups. Viremia levels among dengue patients were notably high for a long period which was assumed to support the spread of the virus by the mosquito vector during epidemic. Phylogenetic analyses of the envelope gene of the epidemic strains revealed close similarity with the strains previously isolated in Myanmar and neighboring countries. DENV-1 dominated the epidemic in 2015 and the serotype (except DENV-3) and genotype distributions were similar in both study sites.

Japanese encephalitis.

Japanese encephalitis (JE) is an inflammation of the central nervous system in humans and animals, specifically horses and cattle. The disease, which can sometimes be fatal, is caused by the flavivirus Japanese encephalitis virus (JEV), of which there are five genotypes (genotypes 1, 2, 3, 4 and 5). The transmission cycle of the virus involves pigs and wild birds as virus amplifiers and mosquitoes as vectors for transferring the virus between amplifying hosts and to dead- end hosts, i.e. humans, horses and cattle. In horses and cattle the disease is usually asymptomatic, but when clinical signs do occur they include fever, decreased appetite, frothing at the mouth, rigidity of the legs and recumbency, and neurological signs, such as convulsive fits, circling, marked depression and disordered consciousness. In pigs, it can cause abortion and stillbirths. At present, the virus is detected in a wide area covering eastern and southern Asia, Indonesia, northern Australia, Papua New Guinea and Pakistan. JEV RNA has also been detected in Italy, first in dead birds in 1997 and 2000 and then in mosquitoes in 2010. Genotype shift, i.e. a change of genotype from genotype 3 to genotype 1, has occurred in some countries, namely Japan, South Korea, Chinese Taipei and Vietnam. Laboratory methods are available for confirming the causative agent of the disease. There are control measures to prevent or minimise infection and, among them, vaccination is one of the most important and one which should be adopted in endemic and epidemic areas.

Silibinin attenuates amyloid beta(25-35) peptide-induced memory impairments: implication of inducible nitric-oxide synthase and tumor necrosis factor-alpha in mice.

In Alzheimer's disease (AD), the deposition of amyloid peptides is invariably associated with oxidative stress and inflammatory responses. Silibinin (silybin), a flavonoid derived from the herb milk thistle, has potent anti-inflammatory and antioxidant activities. However, it remains unclear whether silibinin improves amyloid beta (Abeta) peptide-induced neurotoxicity. In this study, we examined the effect of silibinin on the fear-conditioning memory deficits, inflammatory response, and oxidative stress induced by the intracerebroventricular injection of Abeta peptide(25-35) (Abeta(25-35)) in mice. Mice were treated with silibinin (2, 20, and 200 mg/kg p.o., once a day for 8 days) from the day of the Abeta(25-35) injection (day 0). Memory function was evaluated in cued and contextual fear-conditioning tests (day 6). Nitrotyrosine levels in the hippocampus and amygdala were examined (day 8). The mRNA expression of inducible nitric-oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) in the hippocampus and amygdala was measured 2 h after the Abeta(25-35) injection. We found that silibinin significantly attenuated memory deficits caused by Abeta(25-35) in the cued and contextual fear-conditioning test. Silibinin significantly inhibited the increase in nitrotyrosine levels in the hippocampus and amygdala induced by Abeta(25-35). Nitrotyrosine levels in these regions were negatively correlated with memory performance. Moreover, real-time RT-PCR revealed that silibinin inhibited the overexpression of iNOS and TNF-alpha mRNA in the hippocampus and amygdala induced by Abeta(25-35). These findings suggest that silibinin (i) attenuates memory impairment through amelioration of oxidative stress and inflammatory response induced by Abeta(25-35) and (ii) may be a potential candidate for an AD medication.

Identification of Piccolo as a regulator of behavioral plasticity and dopamine transporter internalization.

Dopamine transporter (DAT) internalization is a mechanism underlying the decreased dopamine reuptake caused by addictive drugs like methamphetamine (METH). We found that Piccolo, a presynaptic scaffolding protein, was overexpressed in the nucleus accumbens (NAc) of the mice repeatedly administrated with METH. Piccolo downexpression by antisense technique augmented METH-induced behavioral sensitization, conditioned reward and synaptic dopamine accumulation in NAc. Expression of Piccolo C2A domain attenuated METH-induced inhibition of dopamine uptake in PC12 cells expressing human DAT. Consistent with this, it slowed down the accelerated DAT internalization induced by METH, thus maintaining the presentation of plasmalemmal DAT. In immunostaining and structural modeling Piccolo C2A domain displays an unusual feature of sequestering membrane phosphatidylinositol 4,5-bisphosphate, which may underlie its role in modulating DAT internalization. Together, our results indicate that Piccolo upregulation induced by METH represents a homeostatic response in the NAc to excessive dopaminergic transmission. Piccolo C2A domain may act as a cytoskeletal regulator for plasmalemmal DAT internalization, which may underlie its contributions in behavioral plasticity.

Population pharmacokinetic analysis of vancomycin in patients with gram-positive infections and the influence of infectious disease type.

OBJECTIVE: To describe the population pharmacokinetics of vancomycin in patients with gram-positive infections and to investigate the influence of type of infectious disease. METHODS: A two-compartment open model was adopted as a pharmacokinetic model. The nonlinear mixed-effects model was used to analyze the population pharmacokinetic models. RESULTS: We propose one general model and one infectious disease type-specific model. The general model showed that vancomycin clearance (CL) was linearly correlated with estimated creatinine clearance (CL(CR)) when CL(CR) was less than 85 mL/min, as expressed by CL(L/h) = 0.0322 x CL(CR) + 0.32. The distribution volumes of the central and peripheral compartment were different in healthy volunteers and patients with gram-positive infections. The infectious disease type-specific model showed that these differences were more pronounced in patients with pneumonia. CONCLUSION: The population pharmacokinetic parameters of vancomycin obtained here can be used to individualize the dosage of vancomycin in institutions with similar patient population characteristics.

Degradation of PEP-19, a calmodulin-binding protein, by calpain is implicated in neuronal cell death induced by intracellular Ca2+ overload.

Excessive elevation of intracellular Ca2+ levels and, subsequently, hyperactivation of Ca2+/calmodulin-dependent processes might play an important role in the pathologic events following cerebral ischemia. PEP-19 is a neuronally expressed polypeptide that acts as an endogenous negative regulator of calmodulin by inhibiting the association of calmodulin with enzymes and other proteins. The aims of the present study were to investigate the effect of PEP-19 overexpression on cell death triggered by Ca2+ overload and how the polypeptide levels are affected by glutamate-induced excitotoxicity and cerebral ischemia. Expression of PEP-19 in HEK293T cells suppressed calmodulin-dependent signaling and protected against cell death elicited by Ca2+ ionophore. Likewise, primary cortical neurons overexpressing PEP-19 became resistant to glutamate-induced cell death. In immunoprecipitation assay, wild type PEP-19 associated with calmodulin, whereas mutated PEP-19, which contains mutations within the calmodulin binding site of PEP-19, failed to associate with calmodulin. We found that the mutation abrogates both the ability to suppress calmodulin-dependent signaling and to protect cells from death. Additionally, the endogenous PEP-19 levels in neurons were significantly reduced following glutamate exposure, this reduction precedes neuronal cell death and can be blocked by treatment with calpain inhibitors. These data suggest that PEP-19 is a substrate for calpain, and that the decreased PEP-19 levels result from its degradation by calpain. A similar reduction of PEP-19 also occurred in the hippocampus of gerbils subjected to transient global ischemia. In contrast to the reduction in PEP-19, no changes in calmodulin occurred following excitotoxicity, suggesting the loss of negative regulation of calmodulin by PEP-19. Taken together, these results provide evidence that PEP-19 overexpression enhances resistance to Ca2+-mediated cytotoxicity, which might be mediated through calmodulin inhibition, and also raises the possibility that PEP-19 degradation by calpain might produce an aberrant activation of calmodulin functions, which in turn causes neuronal cell death.

Repetitive and compulsive-like behaviors lead to cognitive dysfunction in Disc1Δ2-3/Δ2-3 mice.

Disrupted-in-schizophrenia 1 (Disc1) is a key molecular driver for the biology of mental diseases. In order to investigate its role in brain function, we previously generated mice lacking exons 2 and 3 of Disc1 on a C57BL/6J genetic background (Disc1Δ2-3/Δ2-3 mice), which have a deficiency of the full-length Disc1 protein. In the present study, we examined the role of Disc1 in cognitive function using a touchscreen-based visual discrimination (VD) task in which mice had to discriminate 1 of 2 stimuli simultaneously displayed on the screen and received a liquid reward. Disc1Δ2-3/Δ2-3 mice showed impaired performance in the VD task, and this was mainly attributed to the perseverative response being significantly stronger than that in wild-type (WT) mice. Furthermore, the numbers of marbles buried in the marble burying test and nestlets shredded in the nestlet shredding test by Disc1Δ2-3/Δ2-3 mice were significantly higher than those by WT mice, suggesting perseverative/compulsive behaviors by Disc1Δ2-3/Δ2-3 mice. A treatment with clozapine ameliorated behavioral deficits in the VD and marble burying tasks. c-Fos expression was significantly stronger in the dorsomedial striatum (DMS), but not the dorsolateral striatum (DLS) after the first VD session in Disc1Δ2-3/Δ2-3 mice than in WT mice. The treatment of mice that had previously expressed hM3Dq in the DMS with clozapine-N-oxide (CNO) impaired performance in the VD task. These results suggest that cognitive impairments accompanied by perseverative/compulsive behaviors in Disc1Δ2-3/Δ2-3 mice are associated with hyperactivity of the DMS.

Reinforcing effects of morphine are reduced in tissue plasminogen activator-knockout mice.

Tissue plasminogen activator (tPA) plays a key role in neuroplasticity. We have recently demonstrated that the tPA-plasmin system is involved in the rewarding effects of drugs of abuse by regulating the release of dopamine in the nucleus accumbens. In the present study, we investigated whether tPA is involved in the reinforcing properties of morphine in a paradigm of drug self-administration. Eight-week-old tPA knockout and wild-type control mice were subjected to a single 24-h session of morphine self-administration under a fixed ratio (FR) 2 or a progressive ratio (PR) schedule of reinforcement after eight daily 30-min sessions of nose-poke training. tPA knockout mice responded significantly more often for morphine self-administration in a dose-dependent manner as compared with wild-type control mice. Under the PR schedule of morphine reinforcement, however, tPA knockout mice showed a lower breaking point than wild-type control mice. There was no significant difference in food-reinforced operant behavior, breaking points to food pellets, and saline self-administration between the two genotypes. The increased responding in tPA knockout mice under the FR2 schedule was significantly attenuated by the dopamine D1 receptor antagonist SCH23390 (0.3 mg/kg), whereas SCH23390, at a dose range of 0.03-2.0 mg/kg, demonstrated biphasic effects on morphine self-administration in wild-type control mice. Our findings suggest that the reinforcing effects of morphine are reduced in tPA knockout mice. Modulation of the tPA system in the brain may be a potential target against drugs of abuse.

Phencyclidine and genetic animal models of schizophrenia developed in relation to the glutamate hypothesis.

In humans, phencyclidine (PCP), a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, reproduces a schizophrenia-like psychosis including positive symptoms, negative symptoms and cognitive dysfunction. Thus, PCP-treated animals have been utilized as an animal model of schizophrenia. PCP-treated animals exhibit hyperlocomotion as an index of positive symptoms, and a social behavioral deficit in a social interaction test and enhanced immobility in a forced swimming test as indices of negative symptoms. They also show a sensorimotor gating deficit and cognitive dysfunctions in several learning and memory tests. Some of these behavioral changes endure after withdrawal from repeated PCP treatment. Furthermore, repeated PCP treatment induces some neurochemical and neuronanatomical changes. Recently, genetic approaches based on "the glutamate hypothesis of schizophrenia" have been used to develop animal models of schizophrenia. NMDA receptor subunit zeta1 knockdown, epsilon1 knockout (KO) and zeta1 point mutant mice exhibiting a hypofunction of NMDA receptors show hyperlocomotion, social behavioral deficit, sensorimotor gating deficit or cognitive dysfunction. Forebrain-specific calcineurin KO, neuregulin 1 heterozygous KO and lysophosphatidic acid 1 receptor KO mice can also serve as animal models of schizophrenia. These findings suggest that PCP and genetic animal models would be useful for evaluating novel therapeutic candidates and for confirming pathological mechanisms of schizophrenia.

Extended dipyrrin ligands: candidates for optical metal ion detection under competitive conditions.

Acylhydrazone based extended dipyrrins L1-H and L2-H are introduced as complexometric ppb sensitive metal ion detectors. The binding of lanthanide, transition as well as post-transition metal ions is followed by UV-Vis measurements. The carbohydrate based ligand L2 is water soluble and thus can act as a metal ion sensor in this medium.

Stable neutral radicals of planar N2O2-type dipyrrin platinum complexes: hybrid radicals of the delocalized organic π-orbital and platinum d-orbital.

Neutral radicals of N2O2-dipyrrin platinum complexes were synthesized by the reaction of dipyrrin ligands with PtCl2(cod) and successive one-electron oxidation. The radicals are very stable even under aerobic and ambient conditions. X-ray crystallographic analysis revealed the stacking array of the planar dipyrrin complex moieties. The ESR signals were broadened and significantly downfield shifted. The absorption spectra exhibited NIR bands. These results indicated a delocalized radical character with a contribution by the platinum d-orbital.

Hyperfunction of dopaminergic and serotonergic neuronal systems in mice lacking the NMDA receptor epsilon1 subunit.

NMDA receptors, an ionotropic subtype of glutamate receptors (GluRs) forming high Ca(2+)-permeable cation channels, are composed by assembly of the GluRzeta subunit (NR1) with any one of four GluRepsilon subunits (GluRepsilon1-4; NR2A-D). In the present study, we investigated neuronal functions in mice lacking the GluRepsilon1 subunit. GluRepsilon1 mutant mice exhibited a malfunction of NMDA receptors, as evidenced by alterations of [(3)H]MK-801 binding as well as (45)Ca(2+) uptake through the NMDA receptors. A postmortem brain analysis revealed that both dopamine and serotonin metabolism were increased in the frontal cortex and striatum of GluRepsilon1 mutant mice. The NMDA-stimulated [(3)H]dopamine release from the striatum was increased, whereas [(3)H]GABA release was markedly diminished in GluRepsilon1 mutant mice. When (+)bicuculline, a GABA(A) receptor antagonist, was added to the superfusion buffer, NMDA-stimulated [(3)H]dopamine release was significantly increased in wild-type, but not in the mutant mice. GluRepsilon1 mutant mice exhibited an increased spontaneous locomotor activity in a novel environment and an impairment of latent learning in a water-finding task. Hyperlocomotion in GluRepsilon1 mutant mice was attenuated by treatment with haloperidol and risperidone, both of which are clinically used antipsychotic drugs, at doses that had no effect in wild-type mice. These findings provide evidence that NMDA receptors are involved in the regulation of behavior through the modulation of dopaminergic and serotonergic neuronal systems. In addition, our findings suggest that GluRepsilon1 mutant mice are useful as an animal model of psychosis that is associated with NMDA receptor malfunction and hyperfunction of dopaminergic and serotonergic neuronal systems.

Modest neuropsychological deficits caused by reduced noradrenaline metabolism in mice heterozygous for a mutated tyrosine hydroxylase gene.

Tyrosine hydroxylase (TH) is the initial and rate-limiting enzyme for the biosynthesis of catecholamines that are considered to be involved in a variety of neuropsychiatric functions. Here, we report behavioral and neuropsychological deficits in mice carrying a single mutated allele of the TH gene in which TH activity in tissues is reduced to approximately 40% of the wild-type activity. In the mice heterozygous for the TH mutation, noradrenaline accumulation in brain regions was moderately decreased to 73-80% of the wild-type value. Measurement of extracellular noradrenaline level in the frontal cortex by the microdialysis technique showed a reduction in high K(+)-evoked noradrenaline release in the mutants. The mutant mice displayed impairment in the water-finding task associated with latent learning performance. They also exhibited mild impairment in long-term memory formation in three distinct forms of associative learning, including active avoidance, cued fear conditioning, and conditioned taste aversion. These deficits were restored by the drug-induced stimulation of noradrenergic activity. In contrast, the spatial learning and hippocampal long-term potentiation were normal in the mutants. These results provide genetic evidence that the central noradrenaline system plays an important role in memory formation, particularly in the long-term memory of conditioned learning.

Animal models of Alzheimer's disease and evaluation of anti-dementia drugs.

Alzheimer's disease (AD) is the most common cause of progressive decline of cognitive function in aged humans, and is characterized by the presence of numerous senile plaques and neurofibrillary tangles accompanied by neuronal loss. Some, but not all, of the neuropathological alterations and cognitive impairment in AD can be reproduced genetically and pharmacologically in animals. It should be possible to discover novel drugs that slow the progress or alleviate the clinical symptoms of AD by using these animal models. We review the recent progress in the development of animal models of AD and discuss how to use these model animals to evaluate novel anti-dementia drugs.

The Piccolo Intronic Single Nucleotide Polymorphism rs13438494 Regulates Dopamine and Serotonin Uptake and Shows Associations with Dependence-Like Behavior in Genomic Association Study.

Piccolo (PCLO) inhibits methamphetamine-induced neuropharmacological effects via modulation of dopamine (DA) uptake and regulation of the transport of synaptic vesicles in neuronal cells. Clinical studies have recently suggested that the single nucleotide polymorphism (SNP) rs13438494 in the intron 24 of the PCLO gene is associated with psychiatric disorder, in the meta-analysis of GWAS. Therefore, in this study, we attempted to evaluate the possible role of the PCLO SNP in the mechanisms of uptake of monoamines. To characterize rs13438494 in the PCLO gene, we constructed plasmids carrying either the C or A allele of the SNP and transiently transfected them into SH-SY5Y cells to analyze genetic effects on the splicing of PCLO mRNA. The C and A allele constructs produced different composition of the transcripts, indicating that the intronic SNP does affect the splicing pattern. We also transfected DA and serotonin (5-hydroxytryptamine; 5- HT) transporters into cells and analyzed their uptakes to elucidate the association to psychiatric disorders. In the cells transfected with the C allele, both the DA and 5-HT uptake were enhanced compared to the A allele. We also conducted a clinical study, in order to clarify the genetic associations. PCLO rs13438494 exhibits a relationship with the symptoms of drug dependence or related parameters, such as the age of first exposure to methamphetamine, eating disorders, tobacco dependence and fentanyl requirement. Our findings suggest that rs13438494 is associated with drug abuse and contributes to the pathogenesis of psychiatric disorders via modulation of neurotransmitter turnover.