小胶质细胞M1型极化诱发小鼠焦虑样行为及其对前额叶神经元树突棘密度的影响

The effect of microglia M1 polarization on the anxiety-like behaviors and the density of dendritic spines in the medial prefrontal cortex of mice

目的:焦虑样行为与脑内小胶质细胞M1型极化关系密切,但M1型小胶质细胞如何作用于神经元导致焦虑样行为并不清楚,因此本研究的目的在于观察小鼠脑内小胶质细胞向M1型极化诱发产生焦虑样行为,并探讨其对神经元树突棘的影响。方法:实验组采用脂多糖(lipopolysaccharide,LPS)脑内注射诱导C57小鼠小胶质细胞向M1型极化,通过旷场实验和高架十字迷宫实验观察小鼠的行为学改变;应用免疫荧光染色方法观察实验组和对照组小鼠的内侧前额叶皮质(medial prefrontal cortex,m PFC)内一氧化氮合酶(i NOS)与小胶质细胞标志物iba-1的表达变化,通过Western Blot方法观察实验组和对照组小鼠的m PFC内i NOS、致炎因子白介素1(IL-1)β、肿瘤坏死因子(TNF)-α的表达变化,以i NOS、IL-1β和TNF-α表达量均显著增加作为小胶质细胞向M1型极化的标志。采用高尔基(Golgi)染色观察m PFC内神经元树突棘的变化。结果:(1)实验组小鼠mPFC内小胶质细胞i NOS、IL-1β和TNF-α表达量均升高,提示小胶质细胞向M1型极化。(2)实验组小鼠在旷场内的中央活动距离和中央活动时间比对照组均明显减少(P<0.001);实验组小鼠在高架十字迷宫开臂停留时间比正常组明显减少(P<0.05)。(3)Golgi染色结果显示实验组小鼠mPFC内神经元树突棘的密度相较于对照组增多(P<0.05)。结论:mPFC内小胶质细胞M1型极化可诱发小鼠产生焦虑样行为,并伴随mPFC内神经元树突棘密度增多,提示脑内小胶质细胞M1型极化可诱导焦虑样行为,可能与内侧前额叶皮质锥体神经元树突棘可塑性改变相关。

Objective： Microglia polarization to M1 is closely associated with anxiety, while how M1 microglia induce anxiety through neurons is not fully understood. This study was designed to explore the effect of M1 microglia polarization and its effect on anxiety-like behavior and the density of the neuronal dendritic spine in the media prefrontal cortex （mPFC） of mice. Methods： Lipopolysaecharide （LPS） was injected into unilateral lateral ventricle of the C57 mice. Behavioral tests including open field and elevated plus-maze tests were used to observe the anxiety-like behavior of the mice. Immunofluorescent staining and Western blot were applied to detect the expression of inducible nitric oxide synthase （iNOS） in bilateral medial prefrontal cortex （mPFC） after LPS injection to identify the microglia polarization to M1 phe- notype. Western Blot was also used to detect the expression of proinflammatory cytokines like tumor necrosis （TNF）-α and interlukin-1 （IL-1）β. Golgi staining was used to explore the change of neuronal spine density in mPFC. Results： We clearly found that in open field test, LPS treated mice spent significant less time and ran shorter distances in the cen- ter zone, compared with the control group （P 〈 0. 001 ）. In the elevated plus-maze test, LPS treated mice spent less time in the open arm than the control group （P 〈 0. 05 ）. Accordingly, after the LPS treatment, microglia polarized to M1 type as iNOS expression was upregnlated in microglia. The expression of proinflammatory cytokines IL-1 β and TNF-α were sig- nificantly increased after the treatment with LPS. Golgi staining showed that the spine density of the neurons in the mPFC was significantly increased in LPS group, compared with the control group （P 〈 0. 05）. Conclusion ： These data indica- ted that M1 polarization by LPS may induce anxiety-like behavior in the mice and company with increase in the neuronal spine density in the mPFC of the mice. The present study suggests that microglia polarization may lead to anxiety through their influence on dendritic spine of the neurons in mPFC.