Study Of The Effect And Mechanism Of 4-Methylcatechol On Brain-derived Neurotrophic Factor In Rat Brain After Hippocampus Injury

Brain-derived neurotrophic factor (BDNF) is one of the members of the neurotrophin family of proteins, which include nerve growth factor (NGF), neurotrophin-3 (NT-3), NT-4/5, and NT-6. Widespread expression of BDNF mRNA in neurons of the central nervous system (CNS) suggests important roles for BDNF there. BDNF mRNA expression is evoked in association with various brain insults; However, there are two obstacles against the therapeutic application of BDNF to diseases of the CNS. First, BDNF is a macromolecule that cannot pass through the blood-brain barrier (BBB), making it difficult to deliver BDNF from the periphery to the CNS. Second, BDNF is rapidly incorporated into the liver due to its cationic charge, resulting in a short-term circulation of BDNF in the bloodstream. These drawbacks may force consideration of intraventricular infusion of BDNF as therapy, although this approach involves serious technical and/or ethical problems.A promising approach to use neurotrophic actions for therapeutic purposes is the stimulation of synthesis of neurotrophic factors. 4-Methylcatechol (4-MC), a potent stimulator of NGF synthesis in vivo and in vitro, stimulates regeneration of the sciatic, improve and protect against peripheral neuron diseases, Nitta A. et al, had proved that 4MC increases BDNF content and mRNA expression in cultured brain cells and in rat brain in vivo.Objective:To establish rat hippocampus injury model, investigate the effect of 4MCon BDNF production in the brain of hippocampus injury rats. And obtain evidence on suitable 4MC administration time.Methods:①To establish rat hippocampus injury model, Kainic acid was injected ventricularly to young male Wistar rats at a dose known to induce seizures. Take semi-quantify RT-PCR to investigate the change of BDNF mRNA in the hippocampus of model rats. HE staining was used as a way of observing the morphological change of hippocampus neurons in different areas of hippocampus.②To select the most effective dosage, Different dosage of 4MC were intraperitoneal (i.p.) to the model rats the same detect methods as previous research were taken.③To reveal the time when 4MC started to stimulate BDNF synthesis in vivo. 4MC were i.p. to the model rats at different time spots. Besides the previous detecting methods, the distribution of BDNF protein positive neurons in hippocampus was observed with immunohistochemical staining.Results:①The content of BDNF mRNA changed remarkably. T test proved that there was obvious difference between the control group and hippocampus injury model group(t=3.072,P=0.012),indicating that the injury model had been established successfully;HE staining showed that, pyramidical cells in the model group put scattered disorder, and nucleus shrink firmly and darkly stained, the smooth profile of stratum pyramidale hippocampi cells had disappeared. Indicate that hippocampus injury model had been successfully established.②Different dose (0μg/kg.bwt,10μg/kg.bwt,20μg/kg.bwt,50μg/kg.bwt,150μg/kg.bwt)known to effectively induce NGF synthesis in peripheral tissues, were repeatedly administrated to model rats, dose depended enhancement of BDNF mRNA expression was observed in hippocampus. The same trend can bedetected by HE staining.③Semi-quantity RT-PCR proved that there are obvious differences between model group and control group in every time spot group, indicating that we have successfully established hippocampus injury model (P<0.05),obvious differences can be observed among three time spots.6 hour group shows the highest BDNF/β-actin ratio(0.866±0.312),indicating 6 hours after KA injection , may be the best time for 4MC administration.