代谢性谷氨酸受体在脆性X综合征树突棘发育中的作用

Role of mGluR-Ⅰ for the abnormal morphology of dendritic spine in cultured hippocampal neurons of FMR-1 knockout mice

目的探讨代谢性谷氨酸受体-Ⅰ(mGluR-Ⅰ)在脆性X综合征发病中的作用和mGluR-Ⅰ拮抗剂治疗的机制。方法培养FMR-1基因敲除小鼠(KO)和野生(WT)小鼠海马神经元,给予mGluR-Ⅰ激动剂和拮抗剂进行处理,Western-Blot法检测处理前后脆性X智力低下蛋白(FMRP)和微管相关蛋白1B(MAP1B),共聚焦显微镜观察树突棘的长度和密度。结果KO小鼠海马神经元树突棘长度(1·32±0·79)和MAP1B含量显著高于WT小鼠(1·00±0·62)。mGluR-Ⅰ激动剂干预后,KO和WT小鼠树突棘长度(1·60±0·88和1·33±0·80)和MAP1B均显著增加。mGluR-Ⅰ拮抗后,KO小鼠树突棘长度(0·95±0·57)和MAP1B均显著降低,在WT小鼠未见明显变化。结论脆性X综合征模型鼠由于缺乏FMRP的负性调节作用,导致mGluR-Ⅰ激活的MAP1B过度表达,可能是树突棘异常的主要原因。mGluR-Ⅰ拮抗剂逆转树突棘缺陷可能参与治疗机制。

Objective To study the role of metabotropic glutamate receptor- Ⅰ （ mGluR- Ⅰ ） for the abnormal morphology of dendritic spine and microtubule-associated protein 1B （MAP1B） expression in cultured hippocampal neurons of FMR-Ⅰ knockout mice. Methods The cultured hippocampal neurons from FMR-1 gene knockout （ FMR -/- ） and their wild type （ FMR ＋/＋ ） newborn mice were treated with mGluR- Ⅰ agonist （ S ）-3, 5-Dihydroxyphenylglycine （ 100 μmol/L ） and antagonist N-Phenyl-7- （hydroxyimino） cyclopropa-[ b] chromen-1a-carboxamide （ 30 μmol/L） on the 7^th day after plating, The cultures were harvested to determine the level of fragile X mental retardation protein （FMRP） and MAP1B using Western Blot and labeled with the fluorescent marker DiI for the observation of the length and density of dendritic spine using confocal microscopy. Results The length of spine was significantly longer in FMR - / - neurons （ 1.32 ± 0. 79 ） compared to FMR ＋ / ＋ neurons （ 1.00 ± 0. 62 ）. The dendritic spines were elongated and the expression of MAP1B were increased in both FMR - / - and FMR ＋ / ＋ groups （ 1.60 ± 0. 88 and 1.33 ± 0. 80, respectively） treated with the mGluR- Ⅰ agonist （ P 〈 0. 01 ） . The length of dendritic spines and the level of MAP1B were decreased in FMR -/- groups （0. 95 ±0. 57, P 〈0. 01 ） and were not significant changed in FMR ＋/＋ group after treated with antagonist. No significant difference was found in density of dendritic spine between FMR -/- and FMR ＋/＋ groups, treated with neither mGluR-Ⅰ agonist nor antagonist. Conclusions The mGluR-dependent MAP1B synthesis might be exaggerated in the absence of FMRP, which may contribute to increased long dendritic spines in fragile X syndrome, mGluR antagonists reversed the dendritic spine defect, which is considered as a potential treatment for the neurological and psychiatric symptoms.