Experimental infection of mice with Borna disease virus (BDV): replication and distribution of the virus after intracerebral infection

To develop an animal model resembling natural asymptomatic Borna disease virus (BDV) infections, BDV He/80 rat brain homogenate was passaged four times in adult SJL/J mice. Within 12 months of observation, mice did not develop overt signs of disease. Nucleotide sequencing of the rat isolate and the mouse isolates at the fourth passage revealed no difference in the deduced amino acids. Viral RNA was found in brain, heart, kidney, lung, liver, and urinary bladder. Infectious virus was isolated from brain, but also from heart and lung tissue. Immunohistochemically, BDV was demonstrated in nerves in the abdominal cavity, ganglion coeliacum, and adrenal glands, but not in organ parenchyma. Occasionally, viral RNA was detected in mononuclear blood cells.     

Amantadine in depressive patients with Borna disease virus (BDV) infection: an open trial

Objective: Originally introduced into pharmacotherapy as an antiviral compound, amantadine was shown to also have multiple pharmacological effects on the central nervous system. In addition, only a few studies reported on certain antidepressive properties of amantadine. This effect was highlighted by the discovery of its antiviral effect on Borna disease virus (BDV), which is hypothesized to be an etiopathogenetic factor to subtypes of affective disorders. Therefore, the therapeutical use of amantadine in BDV-infected depressive patients was investigated.

Methods: In this open trial, amantadine was added to antidepressive and/or mood-stabilizing compounds treating BDV-infected depressed patients (n=25) with bipolar or major depressive disorders. Amantadine was given twice a day (100–300 mg/day) for a mean of 11 weeks. Antidepressive treatment response was measured on the Hamilton rating scale for depression (HAM-D) and/or with an operationalized diagnostic criteria system (OPCRIT; version 3.31). Virological response was measured by expression of BDV infection parameters in blood samples.

Results: The overall response rate of the amantadine augmentation in the BDV-infected patients with regard to depressive symptoms was 68% after a mean of 2.9 weeks of treatment. Bipolar I patients improved faster and did not show any following hypomania. In addition, the decrease of depression tended to correspond with the decrease in viral activity.

Conclusion: Amantadine appears to show a remarkable antidepressive efficacy in BDV-infected depressive patients. The antidepressive effect in this open trial appeared to be comparable to standard antidepressives, possibly being a result of its antiviral effect against BDV as a potentially relevant etiopathogenetic factor in these disorders.     

Clinical investigation of the relationship between Borna disease virus (BDV) infection and schizophrenia in 67 patients in Japan

The relationship between Borna disease virus (BDV) infection and schizophrenia in the clinical time course was investigated. By nested reverse-transcribed polymerase chain reaction (RT-PCR) and Western blotting, BDV-specific RNA and anti-BDV antibodies were examined in the EDTA-treated blood from 67 schizophrenic patients (according to DSM-III-R) in Japan. A significantly higher proportion (45%) of anti-BDV antibody and/ or BDV RNA carriers were found among these 67 schizophrenic patients than in 26 controls (0%). There were no apparent associations of BDV infection with age, age at onset, period of hospitalization, accompanying somatic diseases, a past history of tuberculosis, a history of transfusion, a family history, or doses of psychotropic drugs. It is possible that, at least, BDV infection in schizophrenic patients may not be a nosocomial (hospital-acquired) infection, although the route of BDV infection in humans remains unidentified. More studies on the relationship between BDV infection and clinical psychosomatic features should be performed in order to elucidate the pathogenesis of schizophrenia.     

Neonatal Borna disease virus infection (BDV)-induced damage to the cerebellum is associated with sensorimotor deficits in developing Lewis rats

Neonatal Borna disease virus (BDV) infection of the brain produces developmental damage to the cerebellum in Lewis rats, with minimal classical inflammatory responses. In the present study, we assessed the consequences of this damage by measuring motor coordination and postural skills in developing (postnatal days 4 to 30) Lewis rats that were neonatally infected with BDV. Neonatal BDV infection-induced motor impairments were selective and correlated with the time course of BDV damage to cerebellar development. BDV-induced motor deficits were not seen until the end of postnatal week 2. By postnatal week 3, BDV-infected rats had deficits in negative geotropism, fore- and hind limb placing and grasping. BDV-infected rats also exhibited deficits in the ability to hold on to a bar and to cross a suspended bar. Neonatal BDV infection induced impairments in the acoustic startle response. Compared to controls, neonatally BDV-infected rats exhibited attenuated habituation of the acoustic startle at postnatal day (PND) 23 and decreased startle responsiveness at PND 30. Prepulse inhibition of the acoustic startle remained unaltered in BDV-infected rats. The data demonstrate that neonatal BDV brain infection of rats can be a valuable animal model system for studying the relationship between abnormal brain development and resultant behavioral deficits. Further studies of this model may elucidate specific pathogenic mechanisms that that may have implications in the study of neurodevelopmental human disorders.     

Developmental alterations in serotoninergic neurotransmission in Borna disease virus (BDV)-infected rats: A multidisciplinary analysis

Neonatal Borna disease virus (BDV) infection of the rat brain serves as a valuable model for studying the pathogenesis of neurodevelopmental abnormalities following early brain injury. Previous experiments have demonstrated significant alterations in regional tissue content of serotonin (5-HT) in neonatally BDV-infected Lewis rats. The present study sought to provide more insights into postnatal virus-associated alterations in 5-HT neurotransmission by evaluating the density of 5-HT1a receptors in the hippocampus and 5-HT2a receptors in the cortex, regional 5-HT tissue concentrations, behavioral responses to a 5-HT agonist, quipazine, and numbers of neurons in specific subfields of the hippocampus on days 7, 14, and 30 after neonatal BDV infection in Lewis rats. Neonatal BDV infection was found to be associated with a gradual increase in the density of 5-HT2a and 5-HT1a postsynaptic receptors followed by an elevation of 5-HT contents at both the levels of synaptic terminals (i.e., cortex and hippocampus) and cell bodies (i.e., raphe nuclei). In addition, there was an enhanced behavioral response to quipazine. Virus-associated neurochemical and behavioral changes were accompanied by a decline in the number of neurons in the dentate gyrus and in the CA1 field of the hippocampus. No change in the number of neurons in the CA3/2 field of the hippocampus was observed. The present pattern of BDV-associated alterations in 5-HT brain system along with available data from other laboratories suggest that BDV might compromise axonal transport and/or release of 5-HT, resulting in decreased 5-HT neurotransmission.     

Wild-type and attenuated influenza virus infection of the neonatal rat brain

Although influenza virus infection of humans has been associated with a wide spectrum of clinical neurological syndromes, the pathogenesis of influenza virus associated central nervous system (CNS) disease in humans remains controversial. To better study influenza virus neuropathogenesis, an animal model of influenza-associated CNS disease using human virus isolates without adaptation to an animal host was developed. This neonatal rat model of influenza virus CNS infection was developed using low-passage human isolates and shows outcomes in specific brain regions, cell types infected, and neuropathological outcomes that parallel the available literature on cases of human CNS infection. The degree of virus replication and spread in the rat brain correlated with the strains' neurotoxicity potential for humans. In addition, using sensitive neurobehavioral test paradigms, changes in brain function were found to be associated with areas of virus replication in neurons. These data suggest that further evaluation of this pathogenesis model may provide important information regarding influenza virus neuropathogenesis, and that this model may have possible utility as a preclinical assay for evaluating the neurological safety of new live attenuated influenza virus vaccine strains.     

Cytokine expression in the rat central nervous system following perinatal Borna disease virus infection

Borna disease virus (BDV) causes central nervous system (CNS) disease in several vertebrate species, which is frequently accompanied by behavioral abnormalities. In the adult rat, intracerebral (i.c.) BDV infection leads to immunomediated meningoencephalitis. In contrast, i.c. infection of neonates causes a persistent infection in the absence of overt signs of brain inflammation. These rats (designated PTI-NB) display distinct behavioral and neurodevelopmental abnormalities. However, the molecular mechanisms for these virally induced CNS disturbances are unknown. Cytokines play an important role in CNS function, both under normal physiological and pathological conditions. Astrocytes and microglia are the primary resident cells of the central nervous system with the capacity to produce cytokines. Strong reactive astrocytosis is observed in the PTI-NB rat brain. We have used a ribonuclease protection assay to investigate the mRNA expression levels of proinflammatory cytokines in different brain regions of PTI-NB and control rats. We show here evidence of a chronic upregulation of proinflammatory cytokines interleukin-6, tumor necrosis factor alpha, interleukins-1alpha, and -1beta in the hippocampus and cerebellum of the PTI-NB rat brain. These brain regions exhibited only a very mild and transient immune infiltration. In contrast, in addition to reactive astrocytes, a strong and sustained microgliosis was observed in the PTI-NB rat brains. Our data suggest that CNS resident cells, namely astrocytes and microglia, are the major source of cytokine expression in the PTI-NB rat brain. The possible implications of these findings are discussed.     

Effects of changes in neonatal thyroid status on the development of neuropeptide systems in the rat brain

The effects of neonatal thyroid deficiency or hyperthyroidism on the development of neurones containing certain neuropeptides was examined in the brains of rats killed at two weeks of age. Five brain areas were dissected and extracted for radioimmunoassay measurement of vasoactive intestinal polypeptide (VIP), somatostatin, cholecystokinin octapeptide (CCK), substance P and neurotensin, whilst corresponding immunocytochemical data were obtained from a quantitative morphological analysis of cell bodies in the cingulate cortex. The two methods of analysis did not always agree, but in hypothyroidism both the concentration of VIP and the number of cells containing VIP-like immunoreactivity were significantly decreased in the anterior and posterior cingulate cortex. In contrast to these effects on the late maturing VIP neurones, the earlier developing somatostatin system was relatively unaffected, whilst neuropeptides localized in cortical fibres rather than cell bodies (such as substance P and neurotensin) were found by radioimmunoassay to be elevated. Hyperthyroidism had less marked effects than neonatal thyroidectomy, although the concentration of CCK (but not the number of immunostained cells) was significantly increased in the cingulate cortex. Radioimmunoassay results from three subcortical areas showed a decrease in VIP concentration in the hypothyroid hypothalamus, and in hyperthyroidism significant elevations of VIP in the basal ganglia, somatostatin in the hypothalamus and CCK in the hippocampus. It appears that in the brain areas studied thyroid disorders result in dis-synchronous shifts in the developmental patterns of the different neuropeptides, and that the effects of thyroid hormone on peptides as on other transmitters are critically dependent on the developmental profile of the system in question.     

Serotoninergic development in the postnatal rat brain

Summary Various characteristics of the developing serotoninergic system in the brain of rats aged 1 to 28 days were studied biochemically.The levels of the precursor amino acid tryptophan showed a maximal increase in the blood, brain and cerebrospinal fluid (CSF) during the 7th and 10th postnatal days. The development of tryptophan hydroxylase activity measuredin vivo by means of 5-hydroxytryptophan (5-HTP) accumulation after NSD 1015 was closely related to the 5-hydroxytryptamine (5-HT) levels at the various ages. 5-HTP accumulation and 5-HT levels increased most markedly after the second postnatal week. 5-Hydroxyindoleacetic acid (5-HIAA) levels were found to increase rapidly in the brain but somewhat more slowly in the CSF during the second week of postnatal development. Regional studies of 5-HTP accumulation after NSD 1015, 5-HT and 5-HIAA levels indicated a caudal to rostral way of maturation.The disappearance of 5-HT was measured after inhibition of tryptophan hydroxylase with H 22/54. The half-life generally decreased in the various brain parts with advancing age, and in the younger animals the shortest half-life was found in the most caudal brain parts. At 28 days of age the half-life was similar in all brain parts studied. These results indicate the existence of an adult like nerve impulse flow in the 5-HT neurons in the brain stem region of the newborn rats. The results from this investigation clearly indicate that the maturation of the different biochemical parameters of the 5-HT pathways develop in a caudal to rostral direction.The study also supports the view that tryptophan hydroxylase may be the limiting step in the development of the serotoninergic system.     

Some effects of chronic antidepressant treatments on rat brain monoaminergic systems

Summary A range of established and putative antidepressant therapies were studied for the effect of their long-term administration on two facets of presynaptic monoaminergic functioning in rat brain, namely NE, DA, and 5-HT turnover and alpha₂-adrenoceptor sensitivity. Unless stated otherwise drugs (10 mg/kg) were injected i.p. twice daily for 14 days. ECT (100 mA for 1 s) was applied once daily for 10 days.Changes in turnover were indirectly assessed by measuring levels of metabolites. Brain levels of MHPG-SO₄ were unchanged by chronic amitriptyline, imipramine, nisoxetine (20 mg/kg), nortriptyline, salbutamol (5 mg/kg), and ECT. Amitriptyline elicited a slight, but significant, increase in brain DOPAC content. Brain levels of 5-HIAA were increased by amitriptyline, imipramine, salbutamol, and ECT. An overall view of the results indicates that no common pattern of change was elicited by the range of antidepressant therapies studied.Central alpha₂-adrenoceptor sensitivity was assessed by investigating the effect of various therapies on the ability of clonidine (25g/kg i.p.) to decrease rat brain MHPG-SO₄ content. The clonidine-induced fall was attenuated by desipramine, imipramine, and ECT. Amitriptyline, iprindole, mianserin, nisoxetine, nortriptyline, Org 6582 (10 mg/kg once daily), pargyline (25 mg/kg once daily), salbutamol, and trazodone were ineffective.The following chronic antidepressant therapies were investigated for their effect on rat frontal cortex³H-clonidine binding: amitriptyline, desipramine, imipramine, iprindole, mianserin, nisoxetine, nortriptyline, pargyline, salbutamol, and ECT. Chronic, but not acute, pargyline decreased³H-clonidine binding and this was due to a diminished number of binding sites.The induction of subsensitive presynaptic alpha₂-adrenoceptors in rat brain is not a property common to all forms of antidepressant therapies. Hence it cannot be the fundamental mode of action of antidepressants. No correlation exists between the changes in rat cortical³H-clonidine binding and the observed changes in the sensitivity of central presynaptic alpha₂- adrenoceptors.     

Effects of a new postnatal stress model on monoaminergic neurotransmitters in rat brains

Introduction: Stress and environmental perturbations influence postnatal brain development and may account for the high disability rates of preterm survivors following intensive care treatment. This study aims to investigate the impact of early environmental factors on the monoaminergic neurotransmitter system in the developing rat brain by using an innovative neonatal stress model. Materials and methods: After birth, in the experimental groups newborn rats were separated from their mothers and exposed to different stressful stimuli four times a day on day P0 to P6 for 10 min each. To mimic intensive care treatment, the stress protocol applied environmental factors like bright light, noise, and low temperature alternating with pain and handling stress at day- and night-time in a varying sequence. The non-stressed control mothers and litters were left completely undisturbed until sacrificing on day P7 or P20. Results: Brains of stressed animals revealed significantly higher levels of norepinephrine (NE) and dopamine (DA) as determined by HPLC-ED and electrochemical detection at day P7 as compared to controls. When returned to their mothers’ undisturbed care, juvenile rats at day P20 still showed higher (yet statistically not significant) concentrations of NE and DA in brain. The stressed animals gained less weight with significantly lower body weights at day P7 compared to controls. Their mothers developed various forms of stressed behaviour. Conclusions: A novel animal model for postnatal intensive care stress was established leading to changes in brain monoamine levels of newborn rats, while undisturbed maternal care seems to moderate the stress effects subsequently.     

博尔纳病病毒感染与精神分裂症关系的初步研究

目的:探讨精神分裂症患者发病与博尔纳病病毒(BDV)感染的关系,分析被BDV感染的精神分裂症患者临床特征。方法:用荧光定量巢式逆转录聚合酶链反应(FQ-nRT-PCR)方法检测86例精神分裂症患者(病例组)和84例健康体检者(对照组)外周血单个核细胞(PBMCs)中BDVp24和BDVp40基因片段,用β-肌动蛋白(β-actin)作为内参照,并总结阳性患者的临床特征,病例组采用阳性与阴性症状量表(PANSS)评分。结果:病例组外周血标本BDV p24基因片段检出率11.6%(10/86),BDV p40基因片段检出率14.0%(12/86),拷贝数均102kb/μl。对照组BDV p24、BDV p40基因片段均为阴性(0%,0/84),两组阳性率差异有统计学意义(P均0.05)。BDV p24和BDV p40基因片段均为阳性的标本测序后,与BDV标准病毒株V和马源的BDV病毒株H1766序列比较同源性分别为96.35%和98.85%。在4个位点出现基因突变(nt1649 T→C、nt1656 G→A、nt1670 C→T和nt1676 C→T)。该目的基因片段与马源的BDV病毒株亲缘关系最近。BDV阳性患者精神症状主要以幻觉、妄想为主。结论:精神分裂症发病与BDV感染有一定的相关性,BDV阳性患者以阳性精神症状为主要临床特征。     

Neonatal Borna disease virus infection in the rat causes a loss of Purkinje cells in the cerebellum.

Viral insults that occur during early postnatal periods, can affect neuronal systems which exhibit significant postnatal development, such as the cerebral cortex and cerebellum. Borna disease virus (BDV) is a single-strand RNA virus which replicates in the nervous system of many species after experimental inoculation and causes acute neurological disease. Neonatal rats infected with BDV do not mount an aggressive response to the virus like their adult counterparts, but instead develop a persistent BDV infection with less overt clinical sequelae. Recently, the cerebellum, a neural structure associated with regulation of motor behavior, and perhaps with higher cognitive functions, has been demonstrated to be a target of neonatal BDV infections in rats (Bautista et al, 1995). In the present study neonatal rats were infected with BDV and their cerebella were analyzed histologically and immunohistochemically at 7 months of age. The cerebella of infected animals were reduced in size but normal foliation and laminar organization was present. However, as visualized with immunohistochemistry for the Purkinje cell-specific antigen calbindin, there were numerous gaps within the Purkinje cell layer and in the molecular layer which contains the Purkinje cell dendritic trees. We estimated the number of Purkinje cells and found there was an approximately 75% loss of PC in adult rats neonatally infected with BDV. These results suggest that neonatal BDV infection may either (1) target the PC and cause the death of these cells directly or (2) acts indirectly by triggering an immune response which is then responsible for the loss of these cells.     

Effects of neonatal bilateral eye enucleation on postnatal development of the monoamines in posterior thalamus of the rat

Summary Levels of dopamine (DA), noradrenaline (NA) and 5-hydroxytryptamine (serotonin, 5-HT) and their metabolites, and the activities of tyrosine hydroxylase (TH), tryptophan hydroxylase (TPH) and monoamine oxidase A and B (MAO-A and MAO-B) have been determined in the rat posterior thalamus after enucleation during postnatal development. DA and 5-HT turnover rate have been measured as 3,4-dihydroxyphenylalanine (DOPA) and 5-hydroxytryptophan (5-HTP) accumulation rates after central decarboxylase inhibition by 3-hydroxybenzylhydrazine (NSD-1015). The major changes were an increase in noradrenergic and serotoninergic metabolism in enucleated animals compared with control animals. A decrease of the MAO-A to MAO-B ratio during postnatal development was found.Abbreviations DA dopamine - DOPAC 3,4-dihydroxyphenylacetic acid - HVA homovanillic acid - DOPA 3,4-dihydroxyphenylalanine - NA noradrenaline - MHPG 3-methoxy-4-hydroxyphenylgly-col - NMN normetanephrine - TRP tryptophan - 5-HTP 5-hydroxytryptophan - 5-HT serotonin - 5-HIAA 5-hydroxy-3-indolacetic acid - TH tyrosine hydroxylase - TPH tryptophan hydroxylase - MAO-A monoamine oxidase-A - MAO-B monoamine oxidase-B Recipients of fellowships from I.I.E. del FISS Reincorporación de doctores y tecnólogos del M.E.C., and Perfeccionamiento de doctores y tecnólogos del MEC, Spain     

重庆地区抑郁症患者博尔纳病病毒感染的分子生物学研究

背景博尔纳病病毒(Borna disease virus,BDV)是一种高度嗜神经的RNA病毒,是人畜共患病博尔纳病(Borna disease,BD)的病原体,可引起从鸟到灵长类的多种动物的中枢神经系统感染[1],表现为以中枢神经系统功能障碍为特征的BD。近年研究发现BDV感染与一些神经精神疾病的发病有关,尤其是精神疾病。但有关BDV感染与抑郁症(depressive disorder,DD)发病之间的关系,目前国内外研究尚有争议。本研究从分子生物学角度进一步探讨BDV与DD发病之间的关系。方法采用巢式逆转录酶聚合酶链反应(nRT-PCR)结合荧光定量聚合酶链反应(FQ-PCR)检测了60例DD患者和120名健康人外周血单个核细胞(PBMCs)中BDV p24基因片段,对FQ-PCR阳性产物进行克隆和基因序列测定,测序结果与人和动物来源的BDV分离株以及标准株Strain V和He/80进行序列比较。对两组阳性率进行Fisher精确概率检验。结果DD组BDV p24基因片段阳性率为5%(3/60);健康组阳性率为0%(0/120)。DD组阳性率高于健康组,差异有显著性意义(P<0·05)。测序结果为5′-CCCTCCAAGTGGAAACCATCCAGACAGCTCAGCGGTGCGACCACTCCGACAGCATCAGGATTCTTGGCGAGAACATCAA-GATACTG-3′。登陆美国国家生物技术信息中心,证实所获得目的基因片段确系BDV p24基因片断,与人类基因组片段和其他病毒基因组片段无同源性。其与马源的BDV病毒株H1766序列比较亲缘关系最近,同源性为97.68%,在2个位点出现突变(nt1675T→C,nt1678C→T)。与其他国际公认的标准病毒株Strain V和He/80比较,同源性分别为96·51%和95·.35%,碱基互换中局限于T-C、和A→G两种。结论中国的DD患者中存在BDV感染,重庆地区DD的发病可能与BDV感染有关。