Continuous cytosine-b-D-arabinofuranoside infusion reduces ectopic granule cells in adult rat hippocampus with attenuation of spontaneous recurrent seizures following pilocarpine-induced status epilepticus

Brief or prolonged seizures induce various patterns of plasticity. Axonal or dendritic remodelling and development of ectopic granule cells have been described in the hilus and molecular layer of the adult rodent hippocampus. Hippocampal cell proliferation also occurs after seizures. However, whether the seizure-induced cell proliferation plays a pathological or reparative role in the epileptic brain is unknown. In this study, we attempted to suppress the seizure-induced cell proliferation with the antimitotic agent cytosine-b-D-arabinofuranoside (Ara-C) and to examine the development of spontaneous recurrent seizures (SRS). Experimental status epilepticus was induced with pilocarpine, and Ara-C or vehicle alone was infused continuously with an osmotic minipump. SRS were video-monitored. BrdU immunohistochemistry was used for the spatial and temporal analysis of hippocampal cell proliferation, and double labelling with NeuN, calbindin and GFAP antibodies was performed for the differentiation of BrdU-positive cells. Timm staining was also performed for evaluation of mossy fibre sprouting (MFS). With continuous Ara-C infusion, the likelihood of developing SRS was decreased and, during the latent period, the development of ectopic granule cells in the hilus and new glia in the CA1 area was reduced when compared with the vehicle-infused group, while MFS was not altered. The results suggest that the hippocampal cell proliferation plays a pro-epileptogenic role rather than a compensatory role, and that the epileptogenic process may be associated with the generation of new glia in the CA1 area and/or new neurons in the dentate gyrus, particularly the ectopically located hilar granule cells.     

Transient reduction of cerebral blood flow leads to longlasting increase in GABA content in vulnerable structures and decreased susceptibility to bicuculline induced seizures

Summary Rats were exposed for 24 min to bilateral clamping of the common carotid arteries (BCCA) in pentobarbital anaesthesia. The GABA content was measured 24 hours, 48 hours, 4 days, 14 days and 3 months after BCCA. In other groups of rats seizures were elicited by i.p. injection of (+)-bicuculline (3 mg/kg) 24 hours, 48 hours, 4 days, 14 days and 3 months after BCCA. Analysis of the GABA content revealed significant increase compared with controls in the hippocampus, frontal cortex and substantia nigra from 24 hours up to 3 months. Bicuculline treatment induced tonic/clonic seizures and status epilepticus in sham operated animals; these effects were drastically diminished at various time points after BCCA. The present results suggest that BCCA produces a longlasting increase in GABA content and as a consequence protection from bicuculline-induced seizures.     

Cerebral metabolic and circulatory changes in the rat during sustained seizures induced by DL-homocysteine

Sustained, generalized seizure activity was induced in anaesthetized (70% N2O), paralyzed and artifically ventilated rats by i.p. DL-homocysteine thiolactone in a dose of 11 mmol/kg. Epileptic discharges in the EEG were accompanied by marked perturbation of tissue metabolites. There was a fall in phosphocreatine concentration to 40% of control but only moderate changes in adenine nucleotides, a marked rise in lactate concentration, and a pronounced increase in the lactate/pyruvate ratio. Excessive amounts of dihydroxyacetone phosphate (and glyceraldehyde phosphate) accumulated, indicating that depletion of NAD+ occurred. There was marked accumulation of ammonia, glutamine and alanine, and reduction in glutamate and aspartate concentrations. Administration of a subconvulsive dose of homocysteine (7.5 mmol/kg) gave rise to changes in ammonia and amino acids, qualitatively similar to those occurring during seizures. It is concluded that although changes in the metabolites of the energy reserve were mainly caused by the induced seizures, those affecting amino acid concentrations were significantly influenced by accumulation of ammonia, secondary to metabolism of injected homocysteine. Cerebral blood flow (CBF) and oxygen utilization (CMRO2) were measured during sustained seizures. CMRO2 rose to 150% of control, with a corresponding increase in CBF.     

Incidence and risk factors of acute encephalopathy with biphasic seizures in febrile status epilepticus

ObjectiveTo clarify the incidence and risk factors of acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) in pediatric patients with febrile status epilepticus (FSE).MethodsWe retrospectively surveyed patients with FSE (≥20 min and ≥40 min) who were younger than 6 years by mailing a questionnaire to 1123 hospitals in Japan. The survey period was 2 years. We then collected clinical data on patients with prolonged febrile seizures (PFS) ≥40 min and those with AESD, and compared clinical data between the PFS and AESD groups.ResultsThe response rate for the primary survey was 42.3%, and 28.0% of hospitals which had applicable cases responded in the secondary survey. The incidence of AESD was 4.3% in patients with FSE ≥20 min and 7.1% in those with FSE ≥40 min. In the second survey, a total of 548 patients had FSE ≥40 min (AESD group, n = 93; PFS group, n = 455). Univariate analysis revealed significant between-group differences in pH, aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, creatine kinase, NH₃, procalcitonin (PCT), uric acid, blood urea nitrogen, creatinine (Cr), and lactate. Multivariate analysis using stratified values showed that high PCT was an only risk factor for AESD. A prediction score of ≥3 was indicative of AESD, as determined using the following indexes: HCO3^? < 20 mmol/L (1 point), Cl <100 mEq/L (1 point), Cr ≥0.35 mg/dL (1 point), glucose ≥200 mg/dL (1 point), and PCT ≥1.7 pg/mL (2 points). The scoring system had sensitivity of 84.2% and specificity of 81.0%.ConclusionIncidence data and prediction scores for AESD will be useful for future intervention trials for AESD.     

Risk and predictors of early epileptic seizures in acute cerebral venous and sinus thrombosis

We assessed the risk and determined predictors of early epileptic seizures (ES) in patients with acute cerebral venous and sinus thrombosis (CVST). A prospective series of 194 consecutive patients with acute CVST admitted to neurological wards in two German university hospitals was analysed for frequency of ES and in-hospital mortality. Demographic, clinical and radiological characteristics during the acute stage were retrospectively analysed for significant association with ES in univariate and multivariate analyses. During the acute stage, 19 patients (9.8%) died. Early symptomatic seizures were found in 86 patients (44.3%). Status epilepticus occurred in 11 patients (12.8%) of whom four died. Amongst patients with epileptic seizures, mortality was three times higher in those with status than in those without (36.4% and 12%, respectively). In multivariate logistic regression analysis, motor deficit [odds ratio (OR) 5.8; 95% CI 2.98–11.42; P  < 0.001], intracranial haemorrhage (OR 2.8; 95% CI 1.46–5.56; P  = 0.002) and cortical vein thrombosis (OR 2.9; 95% CI 1.43–5.96; P  = 0.003) were independent predictors of early epileptic seizures. Status epilepticus was an important source of morbidity and early mortality in patients with CVST in this study. Patients with focal motor deficits, cortical vein thrombosis and intracranial haemorrhage carried the highest risk for ES. Prophylactic antiepileptic treatment may be an option for these patients.     

Local tissue oxygen tension - cytochrome a,a3 redox relationships in rat cerebral cortex in vivo

Simultaneous measurements were made from rat cerebral cortex, in situ, of focal changes in both tissue oxygen tension (ptO2) and the reduction/oxidation ratio of cytochrome c oxidase (cytochrome a,a3) in order to study relationships between oxygen supply and consumption in small regions of tissue. Local ptO2 was measured with polarographic microelectrodes and the redox state of cytochrome a,a3 with a dual wavelength reflectance spectrophotometer. Increased ptO2, produced by respiration of gas mixtures with elevated O2 and/or CO2 content, was accompanied by increased oxidation of cytochrome a,a3. This confirms that cytochrome oxidase is not fully oxidized in focal brain tissue regions in vivo, as it is in mitochondria isolated in vitro. Decreased ptO2 was accompanied by cytochrome a,a3 reduction. The oxidative changes of cytochrome a,a3 with increases in ptO2 were smaller than the reductive changes associated with decreases in ptO2. Curves relating cytochrome a,a3 redox state to ptO2 were qualitatively alike, regardless of the initial ptO2 value from which they were generated. Thus, the reduction level of cytochrome a,a3 varied with ptO2 on a continuum. This consistent relationship demonstrates that changes in mitochondrial redox state provide an index of relative changes in tissue oxygenation in intact neocortex. The results suggest also that local rates of cerebral oxidative metabolism may not always be constant with changes in local ptO2.     

Cerebral metabolic and circulatory changes in the rat during sustained seizures induced bydl-homocysteine

Sustained, generalized seizure activity was induced in anaesthetized (70%N₂O), paralyzed and artificially ventilated rats by i.p.dl-homocysteine thiolactone in a dose of 11 mmol/kg. Epileptic discharges in the EEG were accompanied by marked perturbation of tissue metabolites. There was a fall in phosphocreatine concentration to 40% of control but only moderate changes in adenine nucleotides, a marked rise in lactate concentration, and a pronounced increase in the lactate/pyruvate ratio. Excessive amounts of dihydroxyacetone phosphate (and glyceraldehyde phosphate) accumulated, indicating that depletion of NAD⁺ occurred. There was marked accumulation of ammonia, glutamine and alanine, and reduction in glutamate and aspartate concentrations.Administration of a subconvulsive dose of homocysteine (7.5 mmol/kg) gave rise to changes in ammonia and amino acids, qualitatively similar to those occuring during seizures. It is concluded that although changes in the metabolites of the energy reserve were mainly caused by the induced seizures, those affecting amino acid concentrations were significantly influenced by accumulation of ammonia, secondary to metabolism of injected homocysteine.Cerebral blood flow (CBF) and oxygen utilization (CMR_O2) were measured during sustained seizures. CMR_O2 rose to 150% of control, with a corresponding increase in CBF.     

Hypometabolism precedes limbic atrophy and spontaneous recurrent seizures in a rat model of TLE

Purpose: Temporal hypometabolism on fluorodeoxyglucose positron emission tomography (FDG‐PET) is a common finding in patients with drug‐resistant temporal lobe epilepsy (TLE). The pathophysiology underlying the hypometabolism, including whether it reflects a primary epileptogenic process, or whether it occurs later as result of limbic atrophy or as a result of chronic seizures, remains unknown. This study aimed to investigate the ontologic relationship among limbic atrophy, histological changes, and hypometabolism in rats. Methods: Serial in vivo imaging with FDG‐PET and volumetric magnetic resonance imaging (MRI) was acquired before and during the process of limbic epileptogenesis resulting from kainic acid–induced status epilepticus in the rat. The imaging data were correlated with histologic measures of cell loss, and markers of astrogliosis (glial fibrillary acid protein [GFAP]), synaptogenesis (synaptophysin), glucose transporter 1 (Glut1) and energy metabolism (cytochrome oxidase C), on brains of the animals following the final imaging point. Key Findings: Hippocampal hypometabolism on FDG‐PET was found to be present 24 h following status epilepticus, tending to lessen by 1 week and then become more marked again following the onset of spontaneous seizures. Atrophy of limbic structures was evident from 7 days post‐SE, becoming progressively more marked on serial MRI over subsequent weeks. No relationship was observed between the severity of MRI‐detected atrophy or CA1 pyramidal cell loss and the degree of the hypometabolism on FDG‐PET. However, an inverse relationship was observed between hypometabolism and increased expression of the Glut1 and synaptophysin in the hippocampus. Significance: These findings demonstrate that hypometabolism occurs early in the processes of limbic epileptogenesis and is not merely a consequence of pyramidal cell loss or the progressive atrophy of limbic brain structures that follow. The hypometabolism may reflect cellular mechanisms occurring early during epileptogenesis in addition to any effects of the subsequent recurrent spontaneous seizures.     

NADPH氧化酶及其抑制剂与脑缺血再灌注损伤

缺血性脑血管病中活性氧(ROS)释放量过多,而再灌注后超过身体的抗氧化能力,最终导致脑组织损伤.因此,抗氧化治疗被认为是缺血性脑血管病的治疗方法,但临床实验尚未能将这一概念转化为患者的治疗方案.作为这一概念的转化,目前最主要研究是ROS的来源,而不是ROS本身.在此背景下,NADPH氧化酶(NOX)已被确定为在一般生理条件下和缺血再灌注的脑血管系统中ROS的主要产生者,其主要参与氧化应激、介导自噬、炎症等.抑制NOX可显著减少脑卒中的缺血性损伤,但是NOX家族各亚型的分布和激活机制不同,需要明确其机制及使用抑制剂治疗的重要意义.该文分别对NOX的结构、在脑缺血再灌注损伤中作用机制及抑制剂的研究进展进行综述.     

Rat models of dementia based on reductions in regional glucose metabolism,cerebral blood flow and cytochrome oxidase activity

Summary. Three models are described in rats which attempt to mimic morphological and behavioural pathology of Alzheimers dementia; intracerebroventricular injection of streptozotocin (STZ), permanent bilateral carotid artery occlusion (2VO) and brain mitochondrial cytochrome oxidase inhibition by sodium azide. Learning and memory are impaired within 4 weeks in all models. This probably involves a reduction in cortical and/or hippocampal cholinergic neurotransmission. STZ causes microglial activation and specific damage to myelinated tracts in the fornix through generation of oxidative stress, thereby disrupting connections between the septum and hippocampus. 2VO results in damage to myelin and CA1 cells in hippocampus and in abnormal processing of APP to -amyloid. It is not known if microglial activation and neuronal damage occur after sodium azide administration. Memory and learning can be improved in the STZ and 2VO models by estradiol, melatonin and cholinesterase inhibitors. Antioxidants and neuroprotective agents may also decrease memory deficits by preventing inflammation and neurodegeneration.     

Hippocampal mitochondrial cytochrome C oxidase activity and gene expression in a rat model of chronic cerebral ischemia

The present study established a rat model of chronic cerebral ischemia using bilateral common carotid artery permanent ligation to analyze cytochrome C oxidase activity and mRNA expression in hippocampal mitochondria. Results showed significantly decreased cytochrome C oxidase activity and cytochrome C oxidase II mRNA expression with prolonged ischemia time. Further analysis re-vealed five mitochondrial cytochrome C oxidase II gene mutations, two newly generated mutations, and four absent mutational sites at 1 month after cerebral ischemia, as well as three mitochondrial cytochrome C oxidase III gene mutations, including two newly generating mutations, and one dis-appeared mutational site at 1 month after cerebral ischemia. Results demonstrated that decreased cytochrome C oxidase gene expression and mutations, as well as decreased cytochrome C oxidase activity, resulting in energy dysmetabolism, which has been shown to be involved in the pathological process of ischemic brain injury.     

Suction lesions of the frontal cerebral cortex in the rat induce asymmetrical behavioral and catecholaminergic responses

Suction lesions of the right frontal cerebral cortex in rats induce a period of spontaneous hyperactivity. This hyperactivity, as measured by an increase in running wheel activity begins about one week post-operatively and continues throughout the remainder of a 30-day observation period. The increased activity is accompanied by a bilateral decrease in norepinephrine concentrations in both the ipsilateral and contralateral cortex and locus coeruleus. Identical lesions of the left frontal cerebral cortex produce neither the hyperactivity nor a decrease in norepinephrine concentrations. These experiments have reproduced many of the behavioral and biochemical asymmetries seen after middle cerebral artery ligation; however, suction lesions are both simpler to produce and histologically less variable in their effects.     

Differences in some of the metabolic properties of mitochondrial isolated from cerebral cortex and olfactory bulb of the rat

The metabolic properties of mitochondria from rat cerebral cortex and olfactory bulb were investigated. The pyruvate-supported oxygen uptake rates by olfactory bulb mitochondria were significantly lower than those by cerebrocortical mitochondria. This is consistent with the differences in pyruvate dehydrogenase complex activities between these mitochondrial preparations. Pyruvate dehydrogenase kinase, NAD-linked isocitrate dehydrogenase, and hexokinase activities in olfactory bulb mitochondria were significantly lower than those in cerebrocortical mitochondria. However, NADP-linked isocitrate dehydrogenase, and NAD-linked and NADP-linked glutamate dehydrogenase activities in olfactory bulb mitochondria were significantly higher than those in cerebrocortical mitochondria. The differences between these two mitochondrial preparations in terms of the activities of these energy-metabolizing enzymes reflect the differences detected in the homogenates of these regions.     

A serial analysis of serum aspartate aminotransferase levels in patients with acute encephalopathy with biphasic seizures and late reduced diffusion and prolonged febrile seizure

BackgroundThere are no established biomarkers for diagnosing acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) in the early acute phase, called “the 1st seizure phase”. Based on our clinical experience, we hypothesized that serial examinations of blood levels of aspartate aminotransferase (AST) in children with febrile convulsive status epilepticus (FCSE) revealed higher levels in patients with AESD in the 1st seizure phase than in those with prolonged febrile seizures (PFs).MethodsTo test our presented hypothesis, we retrospectively investigated changes in serum AST in patients with FCSE due to AESD (n = 11) or PFs (n = 27) who were serially examined within 48 h of the onset of convulsions.ResultsThe rate of increase in AST was significantly higher in patients with AESD than in those with PFs. The rate of increase in AST correlated with previously reported scoring systems, i.e., Yokochi and Tottori scores, for the prediction of AESD. A positive correlation between the rate of increase in AST and creatinine levels in the first examination were observed; however, creatinine levels did not significantly differ between the AESD and PFs groups in the first or second examination. Blood levels of pH, ammonia, and sugar in the first examination and C-reactive protein in the second examination were significantly higher in the AESD group than in the PFs group.ConclusionsThe present study revealed that the rate of increase in AST was significantly higher in patients with AESD than in those with PFs. A novel predictive scoring system needs to be established in combination with the rate of increase in AST and reported clinical parameters, which will improve the prognosis of patients with FCSE.     

Changes in midline thalamic recruiting responses in the prefrontal cortex of the rat during the development of chronic limbic seizures

Purpose:   Mesial temporal lobe epilepsy (MTLE) is a common form of epilepsy that affects the limbic system and is associated with decreases in memory and cognitive performance. The medial prefrontal cortex (PC) in rats, which has a role in memory, is associated with and linked anatomically to the limbic system, but it is unknown if and how MTLE affects the PC.
Methods:   We evoked responses in vivo in the PC by electrical stimulation of the mediodorsal (MD) and reuniens (RE) nuclei of the thalamus at several time points following status epilepticus, before and after onset of spontaneous seizures. Kindled animals were used as additional controls for the effect of seizures that were independent of epilepsy.
Results:   Epileptic animals had decreased response amplitudes and significantly reduced recruiting compared to controls, whereas kindled animals showed an increase in both measures. These changes were not associated with neuronal loss in the PC, although there was significant loss in both the MD and RE in the epileptic animals.
Conclusions:   There is a significant reduction in the thalamically induced evoked responses in the PCs of epileptic animals. This finding suggests that physiologic dysfunction in MTLE extends beyond primary limbic circuits into areas without overt neuronal injury.     

脑血管病患者医院感染状况及其相关因素的研究

目的 研究脑血管病患者医院感染状况及其相关因素.方法 对2108例脑血管病患者临床表现、实验室和影像学检查结果进行监测,统计、分析医院感染的发生率、死亡率、感染部位及其相关因素.结果 本组患者发生医院感染726例(34.4%).医院感染组死亡125例(17.2%),无医院感染组死亡112例(8.1%),两组差异有统计学意义(P＜0.01).医院感染发生部位以呼吸道(64.2%)和泌尿道(21.1%)最常见.高龄、住院时间长、脑出血、意识障碍、有侵入性治疗、高血糖及预防性使用抗生素的患者医院感染发生率显著高于年龄较轻、住院时间短、脑梗死、意识清楚、无侵入性治疗、血糖正常及未预防性使用抗生素的患者(均P＜0.01).结论 脑血管病患者医院感染的发生率高,感染患者的死亡率增高.高龄、住院时间长、脑出血、意识障碍、有侵入性治疗、高血糖和预防性使用抗生素是脑血管病患者并发医院感染的常见相关因素.     

Mapping of neuronal networks underlying generalized seizures induced by increasing doses of pentylenetetrazol in the immature and adult rat: a c‐Fos immunohistochemical study

Previous studies from our group have shown that pentylenetetrazol (PTZ)-induced status epilepticus (SE) leads to age-dependent acute and long-term metabolic and circulatory changes in immature rats. In order to define the neural substrates involved in PTZ seizures according to age, the purpose of the present study was to map the areas of cellular activation during seizures of increasing severity in 10-day-old (P10), 21-day-old (P21) and adult rats. Seizures were induced by repetitive injections of subconvulsive doses of PTZ. The total dose received by the animals ranged from 4 to 125 mg/kg. These doses induced a variety of seizure profiles including absence-like, clonic seizures and SE. The cellular activation was measured as the density of c-Fos immunoreactive cells in animals at 2 h after the onset of the seizures. In P10 rats receiving a behaviourally non-active dose of PTZ, c-Fos immunoreactivity appeared only in the amygdala. The dose of 40 mg/kg that induced absence-like seizures led to a weak c-Fos expression in the medial thalamus, some cortical areas and globus pallidus. Clonic seizures reinforced labelling in the previous areas and induced a spread of c-Fos immunoreactivity to other cortical areas, thalamus, hypothalamus and some brainstem nuclei. At that age, only SE led to a widespread and stronger expression of c-Fos which was, however, totally lacking in the midbrain, and remained incomplete in the brainstem and forebrain limbic system, including the hippocampus. In P21 and adult rats, the inactive dose of PTZ induced c-Fos immunoreactivity in thalamus and hypothalamus. With absence-like seizures, c-Fos labelling spread to the cerebral cortex, amygdala, septum and some brainstem regions. With clonic seizures, immunoreactivity was reinforced in all areas already activated by absence-like seizures, and appeared in the striatum, accumbens, brainstem and hippocampus, except in CA1. After SE, c-Fos was strongly expressed in all brain areas. The intensity of c-Fos labelling was higher in most regions of P21 compared to adult rats. These data are in agreement with the immaturity of cellular and synaptic connectivity in P10 rats, the known greater sensitivity of rats to various kinds of seizures during the third week of life and the nature of the neural substrates involved in PTZ seizures.