NR2B受体拮抗剂抑制成年大鼠齿状回新生神经元诱导的长时程增强

NR2B receptor antagonist Ro25-6981 inhibits newborn neuron induced long-term potentiation in the dentate gyrus of adult rats

背景：新生神经元可诱导产生长时程增强，NMDA受体亚基NR2B的激活在成熟神经元诱导的长时程增强中有重要作用，但其对由新生神经元诱导的长时程增强的影响尚未见报道。目的：观察NR2B受体拮抗剂R025-6981在大鼠齿状回新生神经元诱导的长时程增强中的作用。设计、时间及地点：脑片电生理实验，于2007—02／06在山西医科大学神经生物实验室完成。材料：3月龄雄性Wistar大鼠26只，由山西医科大学实验动物中心提供。方法：大鼠麻醉后断头取脑，分离海马，制备400μmol／L脑片。采用细胞外微电极记录技术，于海马齿状回分子层内侧1，3处采用双极钨电极进行低频刺激，获得稳定的刺激曲线后，在高频强直刺激下诱导长时程增强。长时程增强的诱导程序为每串刺激频率100Hz，持续500ms，间隔20s，共4串刺激，记录到兴奋性突触后电位〉1mV以上的脑片用于下述两项实验。①NR2B拮抗剂Ro25—6981阻断人工脑脊液诱导的长时程增强：脑片分为2组，人工脑脊液组持续通以含有95％O2和5％CO2的人工脑脊液，人工脑脊液＋Ro25．6981组在强直刺激前应用3μmol／LNR2B拮抗剂Ro25-6981作用10min，后续步骤同上组。②NR2B拮抗剂Ro25—6981抑制荷包牡丹碱诱导的长时程增强：脑片分为2组，荷包牡丹碱组在强直刺激前给予10μmol／L荷包牡丹碱灌流10min，荷包牡丹碱＋Ro25—6981组在强直刺激前同时给予3μmol／L Ro25．6981和10μmol／L荷包牡丹碱灌流10min。主要观察指标：长时程增强记录结果。结果：①强直刺激后50-60min，人工脑脊液组长时程增强（107．85±1．34）％，人工脑脊液＋Ro25-6981组长时程增强（100．75±2．75）％，两组比较差异有显著性意义（P〈0．05）。②强直刺激后50-60min，荷包牡丹碱组长时程增强（164．67±2．40）％，荷包牡丹碱＋Ro25-6981组长时程增强（147．56±6．63）％，两组比较差异有显著性意义（P〈0．05）。结论：在大鼠海马齿状回区域，NR2B受体拮抗剂Ro25-6981阻断了人工脑脊液诱导的长时程增强，并部分抑制了荷包牡丹碱诱导的长时程增强，提示NR2B受体拮抗剂可以抑制新生神经元诱导的长时程增强。

BACKGROUND： Newborn neurons have been shown to induce long-term potentiation （LTP）. Activation of N-methyI-D-aspartic acid （NMDA） receptor subunit NR2B plays an important role in mature neurons-induced LTP. But there have been no reports addressing on the effects of NR2B activation on newborn neuron-induced LTP. OBJECTIVE： To investigate the effects of NR2B receptor antagonist Ro25-6981 on LTP induced by newborn neurons in adult rat dentate gyrus. DESIGN, TIME AND SETTING： An electrophysiological recording trial was performed at the Department of Neurobiology, Shanxi Medical University from February to June 2007. MATERIALS： Twenty-six male Wistar rats, aged 3 months, were provided by Laboratory Animal Center, Shanxi Medical University. METHODS： Following sacrifice for brain harvesting under anesthesia, the hippocampus was taken to preparation of 400 μ mol/L brain slices. Using extracellular field potential recordings, low-frequency stimulation was performed in the medial molecular layer of dentate gyrus with insulated bipolar tungsten electrodes. After having stable recordings, LTP was induced under high-frequency tetanic stimulation. LTP was induced with a protocol developed previously （4 trains, 500 ms each, 100 Hz within the train, repeated every 20 s）. Only those slices which produced the field excitatory postsynaptic potential of 1 mV or higher in amplitude were accepted for further experiments. （1）NR2B receptor antagonist Ro25-6981 blocked artificial cerebrospinal fluid （ACSF）-induced LTP （ACSF-LTP）： brain slices were divided into 2 groups： ACSF group, in which, slices were continuously perfused using ACSF bubbled with 95% O2 and 5% CO2; ACSF＋ Ro25-6961 group： a 10-minute treatment with 3 μmol/L Ro25-6981 was performed prior to tetanic stimulation, and the remaining procedures were the same as ACSF group. （2） NR2B receptor antagonist Ro25-6981 inhibited bicuculline （BIC）-induced LTP （BIC-LTP）： brain slices were also divided into 2 groups： BIC group： a 10-minute treatment with 10 μ mol/L BIC was performed prior to titanic stimulation; BIC＋ Ro25-6961 group： 3 μ mol/L Ro25-6961 and 10 μmol/L BIC were simultaneously perfused 10 minutes prior to tetanic stimulation. MAIN OUTCOME MEASURES： LTP recording results. RESULTS： （1）After 50-60 minutes of titanic stimulation, LTP was （107.85±1.34）% and （100.75±2.75）% in the ACSF and ACSF＋ Ro25-6981 groups, respectively, with a significant difference between the two groups （P 〈 0.05）. （2） After 50-60 minutes of titanic stimulation, LTP was （164.67±2.40）% and （147.56±6.63）% in the BIC and BIC＋ Ro25-6981 groups, respectively, and a significant difference existed between the two groups （P 〈 0.05）. CONCLUSION： In the rat hippocampal dentate gyrus, NR2B receptor antagonist Ro25-6981 blocked ACSF-LTP and partly inhibited BIC-LTP, indicating that NR2B receptor antagonist can inhibit newborn neuron-induced LTP.