Techniques for functional brain imaging are critical to analyze the information processing of brain and to reveal the advanced functions in brain. These techniques are the hot topics of international research. Great success has been obtained with neuroimaging techniques in the fields of neuroscience research and clinical diagnosis. Existing brain functional imaging such as magnetic resonance imaging (fMRI),positron emission tomography (PET),electroencephalogram (EEG),magnetoencephalography (MEG) and so on,have been successfully used to study brain function. However,these methods have some limitations unavoidably in the temporal or spatial resolution at present. Comparatively,the optical imaging technologies of brain function show their unique charms. Laser speckle imaging (LSI) and intrinsic optical signals imaging (IOSI) stand out because they offer a superior combination of spatial sampling,spatial resolution and temporal resolution; on the other hand,they have no need to use exogenous contrast agents. Great developments also have been obtained in both techniques and applications of brain optical imaging,and they have become powerful tools for in vivo studying functional architecture and pathophysiology in cerebral cortex by monitoring hemodynamics. However,the two optical imaging techniques are confronted with some challenges.

The chemical nature of spinal ganglionic neurons, the peripheral processes of which project divergently to the somatic and visceral areas, has been identified by means of tri-labeling method of combining fluorescein tracing and immunocytochemistry. Ten rats were used. First, 2?l of 2% fast blue (FB) were injected into the left coeliac ganglion. Two days later, 2% nuclear yellow (NY) was injected into left 9-11th intercostal nerves(l?l for each). On the 4th day, animal was perfused with 10% formalin in 0.1mol/L phosphate buffer.The left Th9-11 spinal ganglia were removed and cut into sections by cryostat. The sections were observed under fluorescence microscope and photographed. The results showed that there were three kinds of neurons in the spinal ganglia: (1) single FB labeled cells with blue fluorescent cytoplasm accounted for 38.8% of total labeled cells; (2) single NY labeled cells with yellow fluorescent nuclei accounted for 52,7%; (3) FB and NY double labeled cells accounted for 8.5% and mostly were small or medium in size. Then, the double labeled cells-containing sections were further processed by substance P-demonstrating PAP immunocytochemical staining. The immunostain and fluorescent photographs in the same section were compared and identified each other. We have found that the labeling ratio of SP/NY was 1.4%; SP/FB was 7% and SP/NY+FB was 28.8%. Present study has not only identified the convergence of somato-vesceral sensation in spinal ganglia but also detected the chemical nature of these neurons(substance P) for the first time. In addition, this result has provided a morphological basis for the mechanisma of referred pain and somato-visceral reflection.

Objective To investigate the effect of astrocytes activated by TNF? on the relapse of epilepsy. Methods The purified astrocytes of cultured rat hippocampus were stimulated by TNF?.The expression of NF\|? Bp65 was checked by immunocytochemical method and the release of glutamate(Glu) from astrocytes was measured by high performance liquid chromatography(HPLC).The astrocytes conditioned medium(ACM) stimulated by TNF? was collected and injected into lateral ventricle of chronic Coriaria Lactone kindled rats during the interictal period.The changes both in behavior and electroencephalogram(EEG) were recorded.The expression of NF\|? Bp65 and glial fibrillary acidic protein(GFAP) of hippocampus were also studied by immunocytochemical method. Results 1.TNF? could promote the release of Glu by astrocytes and quickly induce the nuclear expression of NF\|? Bp65 in cultured astrocytes.2.After lateral ventricular injection,four class epileptic behaviors were induced,which were confirmed by epileptic charge EEG.At 0\^5?h after injection of ACM,NF ? Bp65 expression in hippocampal cells especially in the CAl area were induced.It reached to the maximal levels at 2\|4?h and returned to control level at 8h.At 1h after injection of ACM,the increase of the number of positive GFAP \|immunoreactivity(IR) cells of hippocampus could be observed.The expression of GFAP reached to the top level at 4h and still maintained at a higher level than the control group until 8h.Conclusion\ Astrocytes activa\|ted by TNF? could induce the relapse of epilepsy by some solube neuro\|active substance.

Objective To explore the relationship between interleukin-1?(IL-1?) and group Ⅱ metabotropic glutamate receptors(mGluR2/3). Methods The rats were randomly divided into five groups:1^cotrol;2^group Glu;3^group IL-1?+Glu;4^group IL-1?;5^group IL-1?+D-AP5+Glu.The behavior of animals was observed.Expression of mGluR2/3 mRNA was assayed with RT-PCR. Results The alteration of behavior showed that the latent period of seizure in group with IL-1?+Glu was shortened significantly and the degree of seizure was also more serious as compared with that in group Glu.Expression of mGluR2/3 mRNA showed marked enhancement in group with IL-1?+Glu and group with Glu compared with control.However,expression of mGluR2 mRNA in group IL-1?+Glu was markedly decreased as compared to that in group Glu;the level of mGluR3 mRNA showed no apparent difference between group IL-1?+Glu and group Glu.Conclusion Present study suggests that IL-1? enhances the degree of seizure induced by glutamate,the mechanism may be due to its direct or indirect effects on down-regulation of expression of mGluR2 mRNA.

Objective To screen the proteins interacting with AMP-activated protein kinase ?2(AMPK?2)in rat brain cDNA library by bacterial two-hybrid system,and to investigate the biological role and the regulatory mechanism of AMPK?2 in brain.Methods The recombinant pBT-AMPK?2 was used as the bait to screen a rat fetal brain cDNA library by bacterial two-hybrid system.The plasmids of positive colonies were extracted and analyzed by DNA sequencing and BLAST search in GenBank.Results Seven proteins binding to AMPK?2 were screened out,including polyubiquitin,small heat shock protein 8,phosphofructokinase,cytochrome C oxidase subunit Ⅰ(COXⅠ),HLA-B-associated transcript 3(BAT3)isoform 1,protein tyrosine phosphatase receptor type D(Ptprd)and islet-brain 1(IB1).Conclusion Polyubiquitin,HSP8,phosphofructokinase,COXⅠ,BAT3 isoform 1,Ptprd and IB1 can interact with AMPK?2 in brain.

The divergent projections in the bulbo-cerebellar system of rats were studied by double fluorescein labeling method. 2 ?1 of 2％ Fast blue (FB)were injected into the cerebellar cortex of left anterior lobe, 32 hours later, 2 ?l of 2％ Nuclear yellow (NY)were injected into the cerebellar cortex of right anteior lobe. The animals were allowed to survive 16 hours thereafter. Intracardiac perfusion were performed with 10％ formalin in 0.1 mol/L phosphate buffer. The cerebellum and medulla oblongata were removed and serial sections at 20 ?m were cut by cryostat. One of every two sections was preserved for fluorescence microscopic observation. The results identified that there was no intereonnection between the anterior lobes of two cerebellar hemispheres and the anterior lobe of cerebellum received afferent projections from much more nuclei of the medulla oblongata than previous description. The FB or NY single labeled and FB/NY double labeled cells were observed in the nucleus reticularis lateralis, subnucleus reticularis ventralis, nucleus raphe obscurus, nucleus prepositus hypoglossi, nucleus cuneatus lateralis, nucleus spinalis nervi trigemini and nucleus olivaris aceessorius dorsalis. The double labeled cells accounted for 23％ of total labeled cells except the nucleus olivaris inferior. The results mentioned above suggested that the cerebellum receive multiple afferent connections originated from the medulla oblongata, and the divergent projections further amplify the information-receiving areas in the cerebellum, and synchronize the activities of both cerebellar hemispheres.

In order to clarify the regulatory mechanism of the neurohormone releasing in the neurohypophysis, the immunohistochemical and chemical lesion method were combined to demonstrate the vasopressin (VP)-and catecholamine (CA)-containing nerve terminals, and their distribution and relationship were observed under electron microscopic level. The results showed that in the rat neurohypophysis there were not only widely distributed VP nerve terminals, but also there were many 6-OHDA induced degenerated nerve endings. The close relationship even synapse-like contacts existed between the CA-ergic endings and pituicytes as well as microglial cells. It was very interesting that the CA-ergic boutons formed axoaxonic synapses with VP-containing boutons. In this case, the CA-bouton was presynaptic element whereas the VP-bouton served as postsynaptic element. The above mentioned results probably provided ultrastructural evidence for the regulatory mechanism of the neurohormone releasing in the neurohypophysis for the first time.

Origin and ultrastructural characteristics of serotonergic fibers of the spinal dorsal horn in the rat have been confirmed by means of a combined method of HRP and immunocytochemistry and immunoelectron microscopic observation. The results showed that serotonergic axonal terminals in the spinal dorsal horn come mainly from nucleus raphe magnus and the ventral part of the reticular formation of medulla oblongata. Serotonin immunoreactive positive structures of the spinal dorsal horn have been found in lamina Ⅰ (marginal zone) and lamina Ⅱ (substantia gelatinosa) as fine myelinated and unmylinated fibers. There were mainly axo-axonic synapses between the labeled and nonlabeled terminals. The labeled terminals were presynaptic or postsynaptic element. Axo-dendritic synapses were rarely found. The non-synaptic releasing figures have not been found. Based on the ultrastructural characteristics the authors suggest that in performing analgesia role the serotonergic system in the spinal dorsal horn might influence directly or indirectly the excitability of interneurons and inhibit directly the nerve impulses of primary afferents by means of synaptic connections instead of non-synaptic releasing manner.

OBJECTIVE To reconstruct the HNP-1 into the J-HNP-1 with a J chain,and explore to set up a mammalian cell expression system which can express and secret J-HNP-1,so that the products could be examined and purified conveniently. METHODS The J-HNP-1 cDNA fragments were produced by recombinant PCR.Then the J-HNP-1 was inserted into the mammalian expression vector pcDNA3.1(-)/Myc-His which had the double marks Myc and 6?His.The recombinant vector rpcDNA3.1(-)/Myc-His /J-HNP-1 was transfected into the COS-7 cell.The J-HNP-1 expression was analyzed at the mRNA and protein level.The germicidal activity of cell culture supernatant and cellular solution protein was assayed. RESULTS By the use of RT-PCR with special primers,a band of 786bp was amplified from COS-7 cells transfected by this recombinant plasmid.Western blot analysis with specific anti-histidines antibody revealed that the cell culture supernatant and cellular solution protein of COS-7 cells transfected by rpcDNA3.1(-)/Myc-His /J-HNP-1 had a strong band with relative molecular mass of about 24?10~3.Antibacterial activity assay showed that obvious bacterial inhibition occurred in both lysate and supernatant of COS-7 cells transfected by rpcDNA3.1(-)/Myc-His /J-HNP-1. CONCLUSIONS The J-HNP-1 recombinant is obtained and inserted into the mammalian expression vector then to be expressed in vitro.The expression product is found to have the anti-bacterial effect in vitro.

OBJECTIVE To recombine J chain gene with HNP-1 into a new germicidal(molecule) J-HNP-1,which can connect with pIgR by J chain,so that by using pIgR as a "bridge" the J-HNP-1(germicidal) peptide can be transported into the epithelial cell of mucous membrane to kill the intracellular microorganisms,then the recombinant is inserted into the mammalian expression system.METHODS The J chain and HNP-1 cDNA were amplified from the plasmids respectively by PCR,then the two cDNA fragments were recombined into J-HNP-1 by recombinant PCR.The J-HNP-1 cDNA fragment was inserted into the(mammalian) expression vector pcDNA3.1(-)/Myc-HisC.RESULTS The J-HNP-1 recombinant was obtained by(connection) of J chain and HNP-1 cDNA by PCR.The recombinant J-HNP-1 cDNA was 786bp.CONCLUSIONS The recombinant J-HNP-1 cDNA and the construction of expression vector are the basis for the new bactericidal peptide production.