Nikethamide affects inspiratory neuron discharge in the nucleus retrofacialis medial region in brain slices from neonatal rats

BACKGROUND： Nikethamide, a respiratory center stimulant, is widely used in China. However, its effects on the central nervous system and medullary respiratory center remain poorly understood. OBJECTIVE： To investigate the influence of nikethamide on inspiratory neuron discharge in the medial region of the nucleus retrofacialis in neonatal rats, based on the observations addressing rhythmic respiratory discharge generated by the basic medullary respiratory center and various respiration neuron discharges in brain slices. DESIGN, TIME AND SETTING： A controlled, observational study utilizing in vitro neuroelectrophysiology was performed at the Department of Physiology in Southern Medical University between September and December in 2007. MATERIALS： Nikethamide was purchased from Sigma, USA; BL-420E biological signal collection and manaclement system was provided by Chengdu TME Technology, China.METHODS： Isolated medulla-spinal cord preparations were collected from neonatal Sprague Dawley rats, aged 1-3 days. Tissues were divided to include the medial region of the nucleus retrofacialis, ventral respiratory, and dorsal respiratory groups. Subsequently, modified Kreb＇s solution and 5 μg/mL nikethamide-containing modified Kreb＇s solution were consecutively perfused into the medial region of the nucleus retrofacialis in neonatal rat brain slices. MAIN OUTCOME MEASURES： Hypoglossal nerve root respiratory-related rhythmic discharge activities and inspiratory neuron discharges were recorded with an adsorption electrode and microelectrode. RESULTS Nikethamide resulted in prolonged inspiratory neuron discharge time, shortened respiratory cycle and expiratory time. Nikethamide intervention resulted in enhanced integral amplitude of some inspiratory neurons with no changes in discharge frequency or increased discharge frequency in remaining inspiratory neurons with no changes in integral amplitude. CONCLUSION： Nikethamide excites inspiratory neurons in the basic rhythmic respiration and medullary respiratory center, in addition to increased inspiratory neuron and neural network excitability.

Autoregressive trispectrum and its slices analysis of magnetorheological damping device

A combined magnetorheological damper combined with rubber spring and magnetorheological damper is addressed.This type of damping device has inherited the merits of rubber spring and the magnetorheological damper.The test damping device is made up of combined magnetorheological damper,amplitude controller,signal collecting device,computer software for dynamic analysis,etc.When a zeromean and non-Gaussian white noise interfere with the device,a time series autoregressive(AR) model is conducted by using the sampled experimental data.Trispectrum and its slices analysis are emerging as a new powerful technique in signal processing,which is put forward for investigating the dynamic characteristics of the magnetorheological vibrant device.The present of trispectrum and its slices analysis change with the variation of controllable working magnetic field of the damper correspondingly.It is indicated that AR trispectrum and its slices analysis methods are feasible and effective for investigation of magnetorheological vibrant device.

Fast Evaluation of Oxidative DNA Damage by Liquid Chromatography-Electrospray Tandem Mass Spectrometry Coupled With Precision-cut Rat Liver Slices

Objective To establish a fast and sensitive method for the detection of 8-hydroxy-2’-deoxyguanosine （8-OHdG） in precision-cut rat liver slices by HPLC-MS/MS and to investigate isoniazid （INH） -induced oxidative DNA damage. Methods Precision-cut liver slices （300 μm） were prepared from male rats, and incubated with INH （0.018 mol/L） for 2 h after 1 h preincubation. DNA in the slices was extracted and digested into free nucleosides at 37℃ . The samples were injected into HPLC-MS/MS after the proteins were removed. The level of oxidative DNA damage was estimated using the ratio of 8-OHdG to deoxyguanosine （dG）. Results The limit of detection of 8-OHdG was 1 ng/mL （S/N=3） and the intra-assay relative standard variation was 3.38% when one transition 284.3/168.4 was used as a quantifier and another two transitions 284.3/140.2, 306.1/190.2 as qualifiers. 8-OHdG and dG were well separated, as indicated by elution at 10.02 and 7.37 min, respectively. INH significantly increased the ratio of 8-OHdG to dG in rat liver slices （P〈0.05）. Conclusion 8-OHdG in precision-cut liver slices could be sensitively determined by HPLC-MS/MS. HPLC-MS/MS coupled with precision-cut tissue slices is a fast and reliable analytical technique to evaluate oxidative DNA damage of target tissues caused by procarcinogens and cytotoxins.

Oligodendrocyte transcription factor 1 mRNA and protein expression in organotypic rat brain slices

Numerous studies have confirmed that oligodendrocyte transcription factor 1 （Olig-1） is vital for myelin repair. However, the effects of hypoxia and ischemia on Olig-1 expression remain unknown. In this study, Olig-1 mRNA and protein expressions were analyzed by in situ hybridization and immunohistochemistry, to determine the expression profile of Olig-1 in rat brain slices exposed to hypoxia and ischemia. Brains were obtained from 2-day-old Sprague-Dawley rats, and sections were randomly assigned to control and hypoxia/ischemia groups. Hematoxylin-eosin staining revealed karyorrhexis and karyopyknosis in cells from the hypoxia/ischemia group. Under electron microscopy, mitochondria swelling and neuropil edema were observed in the hypoxiaJischemia group. Olig-1 mRNA and protein expressions were increased at 1 day after hypoxia and ischemia treatment. These results suggest that in situ hybridization and immunohistochemistry could be used simultaneously to detect mRNA and protein expression in brain slices.

Protective effect of bone marrow-derived mesenchymal stem cells on dopaminergic neurons against 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in rat brain slices

BACKGROUND： To date, the use of bone marrow-derived mesenchymal stem cells （MSCs） for the treatment of Parkinson’s disease have solely focused on in vivo animal models. Because of the number of influencing factors, it has been difficult to determine a consistent outcome. OBJECTIVE： To establish an injury model in brain slices of substantia nigra and striatum using 1-methyl-4-phenylpytidinium ion （MPP＋）, and to investigate the effect of MSCs on dopaminergic neurons following MPP＋ induced damage. DESIGN, TIME AND SETTING： An in vitro, randomized, controlled, animal experiment using I mmunohistochemistry was performed at the Laboratory of the Department of Anatomy, Fudan University between January 2004 and December 2006. MATERIALS： Primary MSC cultures were obtained from femurs and tibias of adult Sprague Dawley rats. Organotypic brain slices were isolated from substantia nigra and striatum of 1-day-old Sprague Dawley rat pups. Monoclonal antibodies for tyrosine hydroxylase （TH, 1：5 000） were from Santa Cruz （USA）; goat anti-rabbit IgG antibodies labeled with FITC were from Boster Company （China）. METHODS： Organotypic brain slices were cultured for 5 days in whole culture medium supplemented with 50% DMEM, 25% equine serum, and 25% Tyrode’s balanced salt solution. The medium was supplemented with 5 μg/mL Ara-C, and the culture was continued for an additional 5 days. The undergrowth of brain slices was discarded at day 10. Eugonic brain slices were cultured with basal media for an additional 7 days. The brain slices were divided into three groups： control, MPP＋ exposure, and co-culture. For the MPP＋ group, MPP＋ （30 μmol/L） was added to the media at day 17 and brain slices were cultured for 4 days, followed by control media. For the co-culture group, the MPP＋ injured brain slices were placed over MSCs in the well and were further cultured for 7 days. MAIN OUTCOME MEASURES： After 28 days in culture, neurite outgrowth was examined in the brain slices under phase-contrast microscopy. The percent of area containing dead cells in each brain slice was calculated with the help of propidium iodide fluorescence. Brain slices were stained with antibodies for TH to indicate the presence of dopaminergic neurons. Transmission electron microscopy was applied to determine the effect of MSCs on neuronal ultrastructure. RESULTS： Massive cell death and neurite breakage was observed in the MPP＋ group. In addition, TH expression was significantly reduced, compared to the control group （P 〈 0.01）. After 7 days in culture with MSCs, the co-culture group presented with less cell damage and reduced neurite breakage, and TH expression was increased. However, these changes were not significantly different from the MPP＋ group （P 〈 0.01）. Electron microscopy revealed reduced ultrastructural injury to cells in the brain slices. However, vacuoles were present in cells, with some autophagic vacuoles. CONCLUSION： Bone marrow-derived MSCs can promote survival of dopaminergic neurons following MPP＋-induced neurotoxicity in co-cultures with substantia nigra and striatum brain slices.

Diagnostic Value of 16 Slices Spiral-CT for Portal Vein Disorders

Summary: The diagnostic value of 16-slices spiral computed tomography (CT) for portal vein disorders was evaluated. Forty-one patients were scanned by the 16-slices spiral-CT. The celiac trunk, portal vein and their branches were reconstructed by volume rendering (VR), multiplanar volume reconstruction (MPVR) and maximum intensity projection (MIP) technique, and the results were compared with digital subtraction angiography (DSA). VR, MPVR and MIP could display celiac trunk, portal vein, inferior vena cava and their branches and extent of portal vein-vena cava shunt, portal vein emboli and the fistula of hepatic artery-portal vein. The results from 16-slices CT were better than DSA and identical with pathologic ones. The vessel three-dimension reconstruction technique of 16-slices spiral CT is valuable for evaluating the portal systemic disorders.

Simultaneous Measurement of Neural Activities of Acute Mouse Hippocampal Slices Using Multi-Electrode Array System and Laser Confocal Calcium Imaging

Recently, non-invasive, real-time and multi-point measurement of neural activities has become possible by using a multi-electrode array (MEA). Another method for multi-point measurement is the fluorescent imaging technique using voltage indicator dyes or calcium indicator dyes. Especially, calcium imaging using fluorescent calcium indicator dyes is often more useful, because they exhibit larger changes in the fluorescence intensity than voltage indicator dyes and their fluorescence changes can be detect easily. Additionally, calcium signals play key roles in the brain function, such as the long-term potentiation (LTP) in the hippocampus, and calcium imaging can be a powerful tool to elucidate the brain function. In this study, we constructed a measurement apparatus combining the MEA system and laser confocal calcium imaging and simultaneously measured electric signals and calcium signals in acute mouse hippocampal slices. The obtained results showed the availability of the present method.

L-Serine Decreases Taurine Concentration in the Extracellular Fluid of Brain Slices

L-Serine is considered a functional amino acid in the central nervous system, and induces sedation and hypnotic effects in some animal models of acute and chronic stress. Accordingly, while L-serine is a candidate anti-stress factor, the central mechanism of L-serine is not clear. The present study clarifies the action of L-serine using acute chick brain slices. We investigated the changes in some extracellular fluid amino acid concentrations in response to L-serine perfusion. Taurine concentration decreased while L-alanine concentration increased following L-serine perfusion. To examine the involvement of the taurine transporter, the effect of L-serine on the taurine concentration in the presence and absence of Na+ was also investigated. Na+ had no effect on taurine concentration induced by L-serine perfusion. These results suggest that L-serine has an ability to promote L-alanine synthesis facilitating the catabolism of taurine. In conclusion, L-serine modifies the metabolism of taurine and L-alanine in the extracellular space in chick brain.

The Application of Cumulants Slices in Fault Diagnosis

Complex third-order cumulant has different definition forms. Different forms have different coupling properties, and the generated complex cumulants slices contain different coupling information of signals. In experiments, using the different definitions, the same coupling method is applied to both specific fault signals and normal signals. Furthermore, complex third-order cumulant slices spectrum is defined, and it is used to analyse the coupling features of normal signals and fault signals. Experiments indicate that the detection accuracy rate on the same fault is not the same with the different coupling method, thus, it provides an alternative method to diagnose the specific fault.

Coordinated Management of 5G Core Slices by MANO and OSS/BSS

To manage and orchestrate Network Slices (NSs) for 5G Core (5GC), the MANO (MANagement and Orchestration) framework is proposed by European Telecommunications Standard Institute (ETSI). In most research testbeds, MANO systems such as Tacker, OSM and ONAP are used to initiate network slices. However, this doesn’t comply with the 3GPP 5G standards as MANO should only be responsible for dynamic management of NSs, and the static management such as provisioning or unprovisioning a network slice should be left to OSS/BSS (Operation/Business Support System). Thus, in our testbed, an integrated architecture was designed in which the management of network slices will be coordinated by both MANO and OSS/BSS. MANO would handle on-boarding, instantiating, scaling and terminating of network slices while OSS/BSS is responsible for static management of slices including provisioning and unprovisioning of network slices. To evaluate our system, it was compared with the management systems equipped with only OSS/BSS or MANO in order to analyze the shortfalls of those systems when used to deploy network slices. Through this analysis, this research confirms the necessity of applying both OSS/BSS and MANO for the coordinated management of 5G core slices as adopted by 3GPP.

Analytical Solutions of Nonlinear Differential Equations in the Mathematical Model for Inactivation of Nitric Oxide by Rat Cerebellar Slices

A mathematical model for the inactivation of nitric oxide by rat cerebellar slices under non-steady state condition has been analyzed. This diffusion-inactivation model was used to estimate the kinetics of NO consumption by the rat cerebellar slices. He’s Homotopy perturbation method is used to solve the first order nonlinear differential equations which describe the concentrations given by net of diffusion and inactivation by the slices. Analytical expressions for the concentration of nitric oxide have been derived for all values of parameters. The obtained analytical results are compared with the simulation results (Matlab/Scilab program) and are found to be in good agreement.

Evaluating Dedicated Slices of Different Configurations in 5G Core

Network slicing is one of the most important concepts in 5G networks. It is enabled by the Network Function Virtualization (NFV) technology to allow a set of Virtual Network Functions (VNFs) to be interconnected to form a Network Service (NS). When network slices are created in 5G, some are shared among different 5G services while the others are dedicated to specific 5G services. The latter are called dedicated slices. Dedicated slices can be constructed with different configurations. In this research, dedicated slices of different configurations in 5G Core were evaluated in order to discover which one would perform better than the others. The performance of three systems would be compared: 1) Free5GC Stage 2 with each dedicated slice consisting of only UPF;2) Free5GC Stage 3 with each dedicated slice consisting of only UPF;3) Free5GC Stage 3 with each dedicated slice consisting of both SMF and UPF in terms of their registration time, response time, throughput, resource cost, and CPU utilization. It is shown that not one of the above systems will always be the best choice;based on the requirements, a specific system may be the best under a specific situation.

A Heterogeneous Virtual Machines Resource Allocation Scheme in Slices Architecture of 5G Edge Datacenter

In the paper,we investigate the heterogeneous resource allocation scheme for virtual machines with slicing technology in the 5G/B5G edge computing environment.In general,the different slices for different task scenarios exist in the same edge layer synchronously.A lot of researches reveal that the virtual machines of different slices indicate strong heterogeneity with different reserved resource granularity.In the condition,the allocation process is a NP hard problem and difficult for the actual demand of the tasks in the strongly heterogeneous environment.Based on the slicing and container concept,we propose the resource allocation scheme named Two-Dimension allocation and correlation placement Scheme(TDACP).The scheme divides the resource allocation and management work into three stages in this paper:In the first stage,it designs reasonably strategy to allocate resources to different task slices according to demand.In the second stage,it establishes an equivalent relationship between the virtual machine reserved resource capacity and the Service-Level Agreement(SLA)of the virtual machine in different slices.In the third stage,it designs a placement optimization strategy to schedule the equivalent virtual machines in the physical servers.Thus,it is able to establish a virtual machine placement strategy with high resource utilization efficiency and low time cost.The simulation results indicate that the proposed scheme is able to suppress the problem of uneven resource allocation which is caused by the pure preemptive scheduling strategy.It adjusts the number of equivalent virtual machines based on the SLA range of system parameter,and reduces the SLA probability of physical servers effectively based on resource utilization time sampling series linear.The scheme is able to guarantee resource allocation and management work orderly and efficiently in the edge datacenter slices.

Transplanted choroidal plexus epithelial cells can integrate with organotypic spinal cord slices into a new system

This study aimed to evaluate the integration of transplanted choroidal plexus epithelial cells with organotypic spinal cord slices.Organotypic spinal cord slices,normally cultured for 6 days,were divided into control group(Ctrl)and transplanted group(T).The choroidal plexus epithelial cells were dissociated and primary cultured(C group).The choroidal plexus epithelial cells cultured for 6–7 days were labeled by 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanineperchlorate(CM-Dil),and were identified by transthyretin(TTR)in immunocytochemistry.They were adjusted to the density of 0.5–1×107/ml,then 2μl cells suspension were transplanted to the spinal cord slices in the T group.The same amount of basal medium was dripped on the spinal cord slices in the Ctrl group.After 14 days of transplantation,the differentiations into neurons and astrocytes,and the synapses were identified by immunofluorescence histochemistry.At the same time,the ratios of cell differentiations and synapses in new system,and the changes of MAPK signaling pathway were tested by western blotting.The choroid plexus epithelial cells were well labeled by CM-Dil and were immune-stained by TTR in immunocytochemistry.The choroid plexus epithelial cells bodies were small when transplanted on the spinal cord slices,but big when transplanted on the polyester membrane inserts.The transplanted cells could differentiate into astrocytes,and possibly differentiate into neurons,and there were a large number of synaptophysin positive vesicles between transplanted cells and organotypic spinal cord slices in immunofluorescence histochemistry.The levels of GFAP,TUB-III and synaptophysin in the T group were higher than which in the Ctrl and C groups in western blotting(P<0.05).And the ratios of p-JNK/JNK and p-P38/P38 in the T group were significantly lower than which in the Ctrl and C groups(P<0.05).But the ratio of p-ERK/ERK in the three groups was of no significant difference.The transplanted choroidal plexus epithelial cells can integrate with organotypic spinal cord slices into a new system.

Preferential Transport and Metabolism of Glucose in Bergmann Glia over Purkinje Cells:A Multiphoton Study of Cerebellar Slices

了解不同类型的细胞如何处理葡萄糖有助于解释能量供应是如何是如何根据大脑能量需求来进行调整的。荧光追踪结合共聚焦显微镜技术已用于研究培养的脑细胞摄取葡萄糖的实时动态过程。本文采用这种技术利用多光子显微镜观察急性制备的大鼠小脑脑片。带荧光的葡萄糖类似物2NBDG和6NBDG在小脑皮质的分子层中的转运速度比其在蒲肯野细胞胞体和颗粒细胞中快若干倍。洗脱游离示踪剂后,可见大部分磷酸化示踪剂都位于Bergmann胶质细胞,用胶质细胞标记物sulforhodamine 101免疫染色后进一步确认这一结果。有效回收荧光光漂白后显示,2NBDG-P可通过Bergmann胶质细胞之间的缝隙连接沿着分子层水平扩散。本文的结果表明在急性小脑切片中,Bergmann胶质细胞对葡萄糖的转运能力和糖酵解率高于蒲肯野细胞若干倍。由于小脑主要由葡萄糖提供能量,蒲肯野神经元被认为比Bergmann胶质细胞更耗能量,这些结果表明,在胶质细胞和神经元之间存在类似乳酸的能量代谢物介导的环路。

Expression of hypoxia-inducible factor 1 alpha and oligodendrocyte lineage gene-1 in cultured brain slices after oxygen-glucose deprivation

Oligodendrocyte lineage gene-1 expressed in oligodendrocytes may trigger the repair of neuronal myelin impairment, and play a crucial role in myelin repair. Hypoxia-inducible factor la, a transcription factor, is of great significance in premature infants with hypoxic-ischemic brain damage There is little evidence of direct regulatory effects of hypoxia-inducible factor le on oligodendrocyte lineage gene-l. In this study, brain slices of Sprague-Dawley rats were cultured and subjected to oxygen-glucose deprivation. Then, slices were transfected with hypoxia-inducible factor la or oligodendrocyte lineage gene-1. The expression levels of hypoxia-inducible factor la and oligodendrocyte lineage gene-1 were significantly up-regulated in rat brains prior to transfection, as detected by immunohistochemical staining. Eight hours after transfection of slices with hypoxia-inducible factor la, oligodendrocyte lineage gene-1 expression was upregulated, and reached a peak 24 hours after transfection. Oligodendrocyte lineage gene-1 transfection induced no significant differences in hypoxia-inducible factor la levels in rat brain tissues with oxygen-glucose deprivation. These experimental findings indicate that hypoxia-inducible factor la can regulate oligodendrocyte lineage gene-1 expression in hypoxic brain tissue, thus repairing the neural impairment.

Biophoton Radiations Induced by Hydrogen Peroxide in Mouse Liver Slices and Hepatocyte Nuclei in Relation to the Biophysical Action Mechanism of Reactive Oxygen Species

Background: Although a large number of studies have confirmed that the different levels of reactive oxygen species (ROS) in cytoplasm and nucleus have effects on cell growth, proliferation, differentiation and apoptosis, the exact mechanism of ROS action is unclear. An important reason is that the production and degradation time of ROS in cells is very short, and therefore it’s difficult to understand the mechanism of action based on the traditional molecular action process through the ROS diffusion and target binding. Methods: The fresh liver tissue slices were prepared and the nuclei of hepatocytes were separated from Kunming mice according to the reported method. Liver tissue slices and hepatocyte nuclei were perfused with extracellular or intracellular fluids containing different concentrations of hydrogen peroxide (H2O2), and real-time imaging monitoring of biophotonic emission was carried out using an ultra-weak biophoton imaging system. Results: The results showed that the continuous perfusion with different concentrations of H2O2 (300, 400 and 500 μM, respectively) resulted in significant increase of biophotonic emissions, presenting a concentration-dependent effect in liver tissue slices and achieving the maximum effect at 400 μM, while the significant enhancement was found after 500 μM treatment on the hepatocyte nuclei. Conclusion: This study suggests that ROS generated in cells may achieve its physiological and pathological effects via biophotonic emissions, which provides a new quantum biological mechanism of ROS, while the detailed clarification requires further research.