| Epilepsy is a common disease harmful to people's health. It's a complicated syndrome that is induced by the abrupt and intermittent bursting activity of the neuron in the brain, and then manifests repeated paroxysm of the dysfunctional brain. In recent years, some research explored that the occurrence of epilepsy is due to the unbalance of the excitatory system and the inhibitory system in epilepsic patient brain. Melatonin(MT)is an indoleamine produced by several organs and tissues especially the pineal gland. Its chemical name is N-acetyl-5-methoxytryptamin. Many studies reported that melatonin can inhibit convulsion and exert a positive effect on the epileptic activity acrosing its membranes receptors. MT can enhance GABA and GABA receptor activity, to enhance the role of inhibition nervous system and reduce the excitatory amino acids. So that MT can play an important role in protect the nenrons in brain. Howere MT competitive membrane receptor antagonist-Luzindole can block the inhibition role and anticonvulsant role of MT. Substance P (SP) is a member of the tachykinin family. Some evidences suggest that substance P can contributes to hippocampal excitability and state epilepsy, mainly by enhanceing glutamate.It combine with the corresponding G protein-coupled receptor -substance P receptor (SPR / NK1). Substance P and SPR exerts a negative effect on epilepsy. At the same time, a considerable amount of researchs found, SPR can express by inhibitory interneurons of the rat hippocampus. All these remind us that SP&SPR participat in the mechanism of anti-epileptid of MT.But untill now, the mechanism of the melatonin on epilepsy is not very clear. The present study is to observe the influence of melatonin on the behavior of rats induced by Coriaria Lactone (CL) and the influence on the expression of SP and SPR in hippocampus of rats, so as to investigate the mechanism of melatonin acting on epilepsy.In our study, firstly, forty healthy male SD rats were divided into 4 groups at random 10 for each group. They were Group A: NS control group;Group B:CL group;Group C:MT+CL group and Group D:Luzindole+ MT +CL group respectively. We observed and recorded the behavior change and SP,SPR were both detected the in the hippocampus by immunohistochemistry staining method SABC. Finally, We use SYBR Green 1 fluorescence PCR to investigate the SP mRNA in hippocampus of rats of each group. And twenty healthy male SD rats were divided into 4 groups at random, 5 for each group. The way of establishment of model is as sme as immunohistochemistry staining method. Praxiology results showed that the Group B and the Group D displayed serious epileptic convulsion and discharge, which the Group A didn't occur, and the Group C showed slightly outbreak. The immunohistochemistry staining method SABC showed the morphology of neuron in hippocampus and its subregion, We can find SP& SPR positive neurons existed in the CA1-CA3 area and dentate gyrus of hippocampus. The CA1-CA3 region pyramidal neurons containing multipolar cells of various sizes are SPR-positive. The SPR strongly expressed in neuron cytolymph and the nucleus was not staining, much neuron fiber was staining darkly. The SP expressed in pyramidal neuron of hippocampus CA1-CA3 and granular cellslayer of dentate gyrus in each group. And SP also strongly expressed in neuron cytolymph and the nucleus was not staining. The change current of the number of SPR-positive cells and SP-positive cells of each group is similar. The SP & SPR level in the MT+CL group was much decreased than that of the CL group and the Luz+ MT +CL group, staining is much lighter, the distinction had statistical significance (P<0.05). There was no obvious difference between the MT+CL group and control group. SYBR Green 1 fluorescence PCR using the methord of 2-△△CT to evaluate the exoression of SP mRNA in rat hippocampus showed that the relative amount of SP mRNA levels in the rat hippocampus. In CL group and Luz+ MT +CL group, the SP mRNA levels were much higher than that of control group (P<0.05). And the SP mRNA level in the MT+CL group was much more decreased than that of the CL group and the Luz+ MT +CL group (P<0.05). So the result of immunohistochemistry and SYBR Green 1 fluorescence PCR is coincidence. Therefore, our research reval that MT indeed can play an inhibiton role in epilepsy procedure, and indeed the SP SRR participate in the mechanism. Possiblely MT binding with the specific G-protein membrane receptor, directly regulae inhibition system to restrain the activity of epilepsy, can also regulate SP/SPR in brain to restrain epilepsy.We can draw a conclusion that MT may play an significantly inhibitory effect in CL-induced seizures of rats, by reducing the excitability of the SP and the SPR level and activity regulating Glu neurons. This study does not only provide a clear experimental basis for the anti-epileptic mechanism of MT, but also opened up a new therapy of MT for seizures. However, we consider, the mechanism of inhition role of MT on SP&SPR is just one of the mechanisms. Other pathways about anti-epilepsy role of MT need further study. |
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