OBJECTIVETo study the relationship between the protection of Ginsenoside(GS) for spinal cells and nitric oxide (NO).

METHODSpinal cells were cultured in vitro, the model of peripheral nerve was established by scarifying the cells, and NO was measured by Griess method.

RESULTNO in injury group was high than that in noninjury group and NO in group cultured by GS was less than that in group cultured by common medium.

CONCLUSIONNO increases when peripheral nerve is injuried, and the protective effect of GS on spinal cells may be through inhibiting NO release.

Animals ; Cells, Cultured ; Fetus ; Ginsenosides ; isolation & purification ; pharmacology ; Motor Neurons ; cytology ; drug effects ; metabolism ; Neurons, Afferent ; cytology ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Nitric Oxide ; metabolism ; Panax ; chemistry ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley ; Spinal Cord ; cytology ; metabolism

AIMIn order to explore the neurobiological mechanism of morphine addiction and treatment methods, the acute and chronic effects of morphine on the intracellular free calcium concentration ([Ca2+]i) in cultured hippocampal neurons were investigated.

METHODSChanges of [Ca2+]i induced by morphine in primarily cultured hippocampal neurons were measured by confocal laser scanning microscopy using Ca(2+) -sensitive dye fluo-4 as the calcium fluorescent probe.

RESULTSMorphine actually induced the increase in [Ca2+]i of hippocampal neurons. This process could be blocked by naltrindole (delta opioid receptor antagonist) pretreatment, but not by CTOP (micro opioid receptor antagonist) pretreatment. Pretreatment of the cells with thapsigargin almost completely blocked morphine-evoked response; while pretreatment of the cells with verapamil partially inhibited this response. After exposure to 100 micromol/L morphine for 24 h, intracellular [Ca2+]i increased and the increase could be intensified after adding 10 micromol/L naloxone to the medium.

CONCLUSIONMorphine induced the release of Ca2+ is mainly from inositol 1, 4, 5-trisphosphate (IP3) sensitive stores in hippocampal neuron of rats through activation of delta2 subtype opioid receptor.

Animals ; Calcium ; metabolism ; Cells, Cultured ; Hippocampus ; cytology ; Male ; Microscopy, Confocal ; Morphine ; pharmacology ; Neurons ; cytology ; drug effects ; Rats ; Rats, Wistar

OBJECTIVETo investigate the changes of mitochondria membrane potential and cytoplasma cytochrome C as the mechanism of neuron apoptosis induced by B(a)P.

METHODSPrimary neurons were dissociated from cerebral cortex of 1 - 3 days old SD rats and cultured with DMEM incubator at 37 degrees C. After 5 days' cultivation, the neurons were added S9 and B(a)P, and the concentrations of treated B(a)P were 0, 10, 20 and 40 micromol/L respectively. After administering of B(a)P, the neurons were cultivated for 40 hours. Apoptosis rate was measured by flow cytometry using Annexin V-FITC and propidium iodide (PI) staining, and the changes in mitochondrial potential (DeltaPsim) were tested with Rhodamine fluorescence (R2123) technique. Preparation of cytosolic extracts by centrifugation. Western blotting analysis was used to evaluate the level of cytochrome C of cytoplasm.

RESULTSThe apoptotic rate of neuron increased in both the middle dose group and the high dose group compared with controls, and had a dose-response tendency with the concentration of B(a)P. Moreover mitochondrial potential decreased in a dose dependent manner. There was a negative correlation between DeltaPsim and the apoptotic rate of neurons (r = -0.763, P < 0.05); Western blotting analysis showed cytoplasmic cytochrome C level increased significantly, which was positively related with neuron apoptosis (r = 0.831, P < 0.01).

CONCLUSIONLoss of mitochondria membrane potential and increase of cytoplasma cytochrome C may be the main cause of neuron apoptosis induced by B(a)P.

Animals ; Apoptosis ; drug effects ; Benzo(a)pyrene ; toxicity ; Cells, Cultured ; Cytochromes c ; metabolism ; Membrane Potential, Mitochondrial ; Mitochondria ; drug effects ; metabolism ; Neurons ; cytology ; drug effects ; Rats ; Rats, Sprague-Dawley

Erythropoietin (EPO) is an acidic glycoprotein that was first detected as a hematopoietic factor and its synthesis is triggered in response to cellular hypoxia-sensing. EPO binds to type I cytokine receptors, which associate with the non-receptor tyrosine kinase Jak2, and thereby activate Stat 5a/5b, Ras/MAPK, and PI3-K/Akt signaling pathways. The recent discovery shows that there is a specific EPO/EPO-receptor system in the central nervous system (CNS), independently of the haematopoietic system. Hypoxia and anemia can up-regulate EPO/EPOR expressions in the CNS. Further studies demonstrate that EPO has substantial neuro-protective effects and acts as a neurotrophic factor on central cholinergic neurons, influencing their differentiation and regeneration. EPO also exerts neuro-protective activities in different models of brain damage in vivo and in vitro, such as hypoxia, cerebral ischaemia and sub-arachnoid haemorrhage. EPO may also be involved in synaptic plasticity via the inhibition or stimulation of various neurotransmitters. Therefore, human recombinant EPO that activate its receptors in the central nervous system might be utilized in the future clinical practice involving neuroprotection and brain repair.
Animals ; Brain ; metabolism ; Cell Differentiation ; drug effects ; Erythropoietin ; metabolism ; pharmacology ; Humans ; Neurons ; cytology ; drug effects ; Neuroprotective Agents ; metabolism ; pharmacology ; Receptors, Erythropoietin ; metabolism

To study the relationship between tau hyperphosphorylation and the function of glutamate transporter okadaic acid (OA), a protein phosphatase inhibitor, 20 ng in a 0.5 microl volume, was injected into the frontal cortex of rat brain and immunostaining was used to observe the phosphorylation of tau protein and the expression of excitatory amino acid transporter 1 (EAAT1) in the brain following the injection. The results showed that (1) the neurons in the center of the injection region displayed cytoplasmic shrinkage, swelling, nuclear pyknosis, and dislocation at the early stage, and necrosis appeared 3 d after the injection. However, most neurons in the peri-injected areas showed normal morphological characters with immuno positive reaction for AT8, a tau phosphorylated marker; (2) morphological analysis showed that tau hyperphosphorylation caused by OA treatment was mainly observed in the axons and dendrites of neuronal cells at 6 h in the cell body at 1 d, which brought about dystrophic neurites and neurofibrillary tangle (NFT)-like pathological changes; (3) the induction of glutamate transporter EAAT1 was observed in the involved areas corresponding to that with AT8 immunopositive staining, and the number of EAAT1-positive staining cells markedly increased at 12 h (P<0.01), peaked at 1 d (P<0.001), then decreased at 3 d following the injection. Combined with a confocal laser scanning microscopic analysis, double fluorescent immunostaining showed that EAAT1 positive staining appeared in neurons as well as astrocytes in the peri-injected areas of the frontal cortex. These results demonstrate that OA increases glutamate transporter EAAT1 expression in neurons while it induces tau hyperphosphorylation. However, the mechanism and significance of the induction of glutamate transporter EAAT1 expression remain to be further elucidated.
Animals ; Astrocytes ; drug effects ; metabolism ; Axons ; drug effects ; metabolism ; Brain ; cytology ; Dendrites ; drug effects ; metabolism ; Excitatory Amino Acid Transporter 1 ; metabolism ; Neurofibrillary Tangles ; pathology ; Neurons ; drug effects ; metabolism ; Okadaic Acid ; pharmacology ; Phosphorylation ; Rats ; tau Proteins ; metabolism

Animals ; Calcium ; metabolism ; Cell Hypoxia ; Cells, Cultured ; Hippocampus ; cytology ; Neurons ; drug effects ; metabolism ; Rats ; Rats, Wistar ; omega-Conotoxins ; pharmacology

Animals ; Apoptosis ; drug effects ; Cell Hypoxia ; Cells, Cultured ; Hippocampus ; cytology ; Neurons ; cytology ; drug effects ; Propofol ; pharmacology ; Proto-Oncogene Proteins c-fos ; metabolism ; Rats

AIMTo explore IL-6 neuroprotection against glutamate-induced neurotoxicity and primary mechanisms involved in this neuroprotection.

METHODSThe cerebellar granule neurons from postnatal 8-day infant rats were chronically exposed to IL-6 for 8 days, and then glutamate stimulated the cultured cerebellar granule neurons for 15 min. Methyl-thiazole-tetrazolium (MTT) assay and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method were used to observe the changes of neuronal vitality and apoptosis, respectively. Laser scanning confocal microscope (LSCM) and reverse transcription-polymerase chain reaction (RT-PCR) were respectively employed to measure dynamic changes of intracellular Ca2+ levels and expression of gp130 mRNA, a 130-kDa intracellular IL-6 signal-transduction protein, in the neurons.

RESULTSThe chronic IL-6 (2.5, 5 and 10 ng/ml) pretreatment of the cultured cerebellar granule neurons remarkably improved the decreased neuronal vitality by glutamate in a concentration-dependent manner. The neuronal apoptosis induced by glutamate was significantly attenuated by the chronic IL-6 pretreatment. The intracellular Ca2+ overload evoked by glutamate was also inhibited by the chronic IL-6 pretreatment. The expression of gp130 mRNA was dramatically lower in the IL-6-pretreated cerebellar granule neurons than in the IL-6-untreated neurons.

CONCLUSIONIL-6 can protect neurons against glutamate-induced exciting neurotoxicity. The mechanism of IL-6 neuroprotection may be closely related to the suppression of glutamate-induced intracellular Ca2+ overload and mediated by gp130 intracellular signal transduction pathways.

Animals ; Cells, Cultured ; Cerebellum ; cytology ; drug effects ; metabolism ; Glutamic Acid ; toxicity ; Interleukin-6 ; pharmacology ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Neurotoxicity Syndromes ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo establish the in vitro model of PGE2 released by hypothalamic neurocytes under rrIL-1beta in vitro interference, and investigate the correlation of the PGE2 content and the effect of the drug effect concentration in the model under the effect of Bupleurum injection.

METHODHypothalamic neurocytes were cultured in vitro, and added with rrIL-1beta (40 microg x L(-1)) stimulation. Cell sap was collected at different time points. ELISA was adopted to determine the content of PGE2 in cell sap collected at different time points. Hypothalamic neurocytes were cultured in vitro, added with rrIL-1beta (40 microg x L(-1)) stimulation and then different concentrations of Bupleurum injection. The changes in the content of PGE2 in cell supernatant were detected by ELISA. An analysis was made on the linear relationship between the sample concentration and the inhibition rate of PGE2.

RESULTThe rrIL-1 cells could stimulate in vitro cultured hypothalamic neurocytes to release PGE2 and reach the peak at 10 h. Bupleurum injection could significantly interfere the release of PGE2 in the in vitro model (P < 0.01, P < 0.05), with a certain linear relationship between the interference effect and the effect concentration of Bupleurum injection (r = 0.911, P < 0.01).

CONCLUSIONThe rrIL-1 cells could stimulate in vitro cultured hypothalamic neurocytes to release PGE2, with a good correlation between the inhibition and generation effects of PGE2 and the drug concentration.

Animals ; Biological Assay ; Bupleurum ; chemistry ; Cells, Cultured ; Dinoprostone ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Female ; Hypothalamus ; cytology ; drug effects ; metabolism ; Neurons ; drug effects ; metabolism ; Rats ; Rats, Sprague-Dawley

AIMTo study the protective effect of Quinacrine(QA) on rat striatum neurons from the injury caused by heat environment treatment, to probe the relationship between cell membrane injury and cellular injury protection, and to seek the possibility of QA as a preventive agent to heat injury.

METHODSPrimary cultured striatum neurons from newborn rats were pretreated with QA at different concentration for 1 h, and then heat-treated at 43 degrees C for another 1 h. Cell necrosis was detected by Trypan blue staining, and apoptosis was evaluated through Activated Caspase-3 dye and TdT dye.

RESULTSHeat treatment effected the survival of striatum neurons and resulted in great number of cell death, which was mainly mediated by cell necrosis process. It was shown that treatment of QA itself had little effect on the survival of striatum neurons, while QA pretreatment decreased cellular necrosis caused by following heat treatment.

CONCLUSIONQA protects striatum neurons from heat environment injury at about 20 pmol/L, and the protection may mediated by reduction of necrosis.

Animals ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cell Death ; drug effects ; Cells, Cultured ; Corpus Striatum ; cytology ; Heat-Shock Response ; Neurons ; drug effects ; Quinacrine ; pharmacology ; Rats ; Rats, Wistar