| Objective To observe the seizure ,the positive serotonergic neuron in raphe nuclei and the dynamics of hippocampal release of serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) in pilocarpine-induced epileptic rats after 5,7-dihydroxytryptamine (5,7-DHT) microinjection in median raphe nucleus (MnR),then investigate the relationship between 5-HT and epilepsy. In the meanwhile,evaluate the epiletic rat 's ability of spatial learning and memory in Morris water maze (MWM),then study the mechanism of learning and memory in pilocarpine-induced epileptic rats.Methods1.Experimental animal and groups:The first part:60 male adult Sprague-Dawly rats were randomly divided into 3 groups:vehicle control group,pilocarpine(PILO) treated group(PILO+NS) and PILO+ 5,7-DHT treated group(PILO+5,7-DHT).In the first part experiment,PILO+NS group was divided into two groups: have status epilepticus(SE) and no SE.2. According to Watson《rat biain stereotaxic atlas》, recording electrode was inserted in right frontal lobe cortex ,microdialysis probe was inserted in right hippocampal and 5,7-DHT was microinjected into MnR(median raphe nucleus) with the help of brain solid positioner.3.Establish pilocarpine-induced epileptic model: intraperitoneal injection PILO(320mg/kg or 30mg/kg).4.Apply vedio electroencephalography (EEG) to observe the rats'seizures and cortex EEG.5.Use Morris water maze to evaluate the rats' spatial learning-memory.6. Make use of immunohistochemisty observing serotonergic neuron in raphe nuclei.7.Apply brain microdialysis technology combined with high-performance liquid chromatography(HPLC)-electrochemistric determination(ECD) to observe the dynamics of hippocampal release of serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA).Results1.Success rate and mortality in pilocarpine-induced epilepsy model: after treatment of 5,7-DHT(PILO+5,7-DHT),the success rate and the mortality were all improved.2.Behavior change:the seizure of the rats cuuld be devided into three phases: acute period, silent period and chronic period after treatment of PILO. Compared with the PILO+SE group, the chronic spontaneous seizures'frequency were increase in PILO+5,7-DHT group(P<0.05),while we did not abserve spontaneous seizures in PILO-SE group.3.EEG:Acute period:spike-wave or Spike-and-slow-wave complex were oberved.Silent period:a-wave was accompanied by a few low sharp-waves or Spike-and-slow-wave complex.Chronic period: sharp-spike-waves, Spike-and-slow-wave complex and multiple Spike-and-slow-wave complex were oberved.4.Morris water maze (MWM) performance: compared with control group,the mean escape latency was prolonged,the number of crossing target was decreased and the retention time in target zone was shortened in PILO+SE group(P<0.05),but there was no significant difference between PILO+SE group and PILO+5,7-DHT group(P>0.05).5.The number of serotonergic neuron in raphe nuclei: compared with the control group ,the number of serotonergic neuron in raphe nuclei was decrease in status epilepticus group (PILO+SE) (P<0.05).Moreover,it's extremely decrease in PILO+5,7-DHT group(P<0.01).6.The content of extracellular 5-HT and 5-HIAA in hippocampus: in PILO+NS group, there was a great increase of 5-HT and 5-HIAA during acute period , no change during silent period and decrease during chronic period when compared with the control group.Compared with control group, there was a great decrease of 5-HT and 5-HIAA during chronic period in PILO+5,7-DHT group. Conclusions1. We have established stable technology of brain microdialysis in pilocarpine-induced epileptic rats,which laid the foundation of observing the dynamics of hippocampal release of 5-HT and 5-HIAA.2. Depletion of serotonin may facility the rats'epileptic seizures.3. Epileptic seizures perhaps is one of the factors causing epileptic rats' spatial learning and memory deficit.But we could not interpret the level of brain 5-HT was relevant with the change mechanism of learning and memory in epiletic rat. |
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