[1]王来法,王雪琴,邓玲,等.小胶质细胞抑制剂Pexidartinib对小鼠遥远场景性恐惧记忆再巩固的影响[J].新乡医学院学报,2023,40(12):1107-1113.[doi:10.7683/xxyxyxb.2023.12.002]
　WANG Laifa,WANG Xueqin,DENG Ling,et al.Effect of microglia inhibitor Pexidartinib on reconsolidation of remote contextual fear memory in mice[J].Journal of Xinxiang Medical University,2023,40(12):1107-1113.[doi:10.7683/xxyxyxb.2023.12.002]
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小胶质细胞抑制剂Pexidartinib对小鼠遥远场景性恐惧记忆再巩固的影响
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《新乡医学院学报》[ISSN:1004-7239/CN:41-1186/R]

卷:
40卷
期数:
2023年12
页码:
1107-1113
栏目:
基础研究
出版日期:
2023-12-05

文章信息/Info

Title:
Effect of microglia inhibitor Pexidartinib on reconsolidation of remote contextual fear memory in mice
作者:
王来法; 王雪琴; 邓玲; 张辉; 崔艳慧
（长沙医学院神经科学与行为学研究中心，湖南　长沙　410219）
Author(s):
WANG Laifa; WANG Xueqin; DENG Ling; ZHANG Hui; CUI Yanhui
(Neuroscience and Behavioral Research Center,Changsha Medical University,Changsha 410219,Hunan Province,China)
关键词:
小胶质细胞; Pexidartinib; 遥远场景性恐惧; 记忆再巩固; 磷酸化溴结构域蛋白 4; 消皮素 D; 混合谱系激酶结构域样蛋白
Keywords:
microglia; Pexidartinib; remote contextual fear; memory reconsolidation; phosphorylated bromodomain-containing protein 4; gasdermin D; mixed lineage kinase domain-like protein
分类号:
R749
DOI:
10.7683/xxyxyxb.2023.12.002
文献标志码:
A
摘要:
目的　探讨小胶质细胞抑制剂Pexidartinib对小鼠遥远场景性恐惧记忆再巩固的影响。
方法　将12只健康C57BL/6雄性小鼠随机分为实验组和对照组，每组6只。2组小鼠均置于场景恐惧反应箱中进行场景性恐惧条件化训练，构建场景性恐惧模型，并记录每次电击后小鼠的僵直时间；7 d后，实验组小鼠给予含Pexidartinib制剂PLX3397的鼠粮喂养，对照组小鼠给予普通鼠粮喂养，直至行为实验结果。场景性恐惧条件化训练后第16天，将小鼠重新放入场景恐惧反应箱中进行恐惧记忆唤起，不给予任何刺激呈现，小鼠自由探索5 min后取出，记录小鼠在此期间的僵直时间。在恐惧记忆唤起后24 h，将小鼠再次置于场景恐惧反应箱中，使其自由探索3 min，并记录其在该时间段所呈现的僵直时间。采用僵直时间百分比来表示小鼠的恐惧反应。行为实验结束后，腹腔注射戊巴比妥钠麻醉小鼠。2组各取3只小鼠迅速开胸暴露心脏，将灌注针自心尖插入左心室，使用4 ℃预冷的40 g·L-1多聚甲醛(pH=7.4)进行灌注，至小鼠全身不自主抽动消失、全身肢体僵硬为止，取脑组织，多聚甲醛溶液固定，置于蔗糖溶液进行沉糖，然后用组织包埋剂包埋，采用免疫组织化学染色法检测2组小鼠海马中小胶质细胞数量。2组其余小鼠迅速断头取脑组织，冰上分离两侧海马组织，采用Western blot法检测小鼠海马组织中磷酸化溴结构域蛋白4 (pBRD4）、消皮素 D（GSDMD）、混合谱系激酶结构域样蛋白（MLKL）蛋白表达。
结果　在场景性恐惧条件化训练阶段，2组小鼠的僵直时间百分比均随足底电击次数的增加而升高（P<0.05），但2组小鼠第1、2、3、4、5次电击后的僵直时间百分比比较差异无统计学意义（P>0.05）。在唤起阶段，2组小鼠的僵直时间百分比比较差异无统计学意义（P>0.05）。在记忆检测阶段，实验组小鼠的僵直时间百分比显著低于对照组（P<0.05）。实验组小鼠海马CA1、CA3和DG 区小胶质细胞数量均显著低于对照组（P<0.05）。实验组小鼠海马组织中pBRD4、GSDMD和MLKL蛋白相对表达量显著低于对照组（P<0.05）。
结论　小胶质细胞抑制剂Pexidartinib能够损伤遥远场景性恐惧记忆的再巩固，其作用机制可能与其抑制小胶质细胞的激活及下调pBRD4、GSDMD和MLKL的表达有关。
Abstract:
Objective　To investigate the effect of microglia inhibitor Pexidartinib on reconsolidation of remote contextual fear memory in mice.
Methods　Twelve healthy C57BL/6 male mice were randomly divided into experimental group and control group,with 6 mice in each group.The mice in the two groups underwent contextual fear conditioning training in the contextual fear response box to establish the contextual fear models,and the freezing time of mice after each footshock was recorded.After 7 days,the mice in the experimental group were fed with food containing Pexidartinib formulation PLX3397,while the mice in the control group were fed with regular food until the end of behavioral experiment.On the 16th day after contextual fear conditioning training,the mice were put back into the contextual fear box for recalling the fear memory without any stimulation.The mice were taken out after 5 minutes of free exploration,and the freezing time of mice during this period was recorded.At 24 h after the fear memory was recalled,the mice were again placed in the contextual fear box,allowing them to explore freely for 3 minutes,and the freezing time of mice during this period was recorded;the fear response of mice was indicated by the percentage of freezing time.After the behavioral experiment,the mice were anesthetized by intraperitoneal injection of pentobarbital sodium,and three mice from the two groups were taken and rapidly opened the chest to expose their hearts,the perfusion needle was inserted into the left ventricle from the tip of heart and then was perfuse with 40 g·L-1 paraformaldehyde (pH=7.4) which precooled at 4 ℃ until the involuntary convulsion disappeared and the body limbs of mice were stiff.The brain tissue of mice was taken and fixed with paraformaldehyde solution,and then placed in sucrose solution for dehydration.The brain tissue of mice was coated with tissue embedding agent.The number of microglial cells in the hippocampus of mice in the two groups was detected by immunohistochemistry.The remaining mice in the two groups were taken and quickly decapitated to obtain brain tissues,and the hippocampus tissues of two sides were separated on ice.The expressions of phosphorylated bromodomain-containing protein 4 (pBRD4),gasdermin D(GSDMD) and mixed lineage kinase domain-like protein(MLKL) in the hippocampus of mice were detected by Western blot.
Results　In the stage of contextual fear conditioning training,the percentage of freezing time of mice in two groups increased with the increase of the number of footshock(P<0.05),but there was no significant difference in the percentage of freezing time of mice after the first,second,third,fourth and fifth footshock between the two groups (P>0.05).There was no significant difference in the percentage of freezing time of mice between the two groups during recall period(P>0.05).The percentage of freezing time of mice in the experimental group was significantly lower than that in the control group at the phase of memory test (P<0.05).The number of microglia in CA1,CA3 and DG regions of hippocampus of mice in the experimental group was significantly lower than that in the control group (P<0.05).The relative expressions of pBRD4,GSDMD and MLKL in hippocampus of mice in the experimental group were significantly lower than those in the control group (P<0.05).
Conclusion　Microglia inhibitor Pexidartinib can injury the reconsolidation of remote contextual fear memory,which may be related to its inhibition of microglial cell activation and the down-regulation of the expressions of pBRD4,GSDMD and MLKL.

参考文献/References:

［1］　AL ABED A S,DUCOURNEAU E G,BOUARAB C,et al.Preventing and treating PTSD-like memory by trauma contextualization［J］.Nat Commun,2020,11(1):4220.
［2］　JOHNSON L R,MCGUIRE J,LAZARUS R,et al.Pavlovian fear memory circuits and phenotype models of PTSD［J］.Neuropharmacology,2012,62(2):638-646.
［3］　SUMNER J A,NISHIMI K M,KOENEN K C,et al.Posttraumatic stress disorder and inflammation:untangling issues of bidirectionality［J］.Biol Psychiatry,2020,87(10):885-897.
［4］　DUCOURNEAU E G,GUETTE C,PERROT D,et al.Brexpiprazole blocks post-traumatic stress disorder-like memory while promoting normal fear memory［J］.Mol Psychiatry,2021,26(7):3018-3033.
［5］　MURKAR A,KENT P,CAYER C,et al.Gastrin-releasing peptide attenuates fear memory reconsolidation［J］.Behav Brain Res,2018,347:255-262.
［6］　HUANG B,ZHU H,ZHOU Y,et al.Unconditioned- and conditioned- stimuli induce differential memory reconsolidation and β-AR-dependent CREB activation［J］.Front Neural Circuits,2017,11:53.
［7］　ALI VAFAEI A,NAZARI M,OMOUMI S,et al.Corticosterone injection into the basolateral amygdala before and after memory reactivation impairs the subsequent expression of fear memory in rats:an interaction of glucocorticoids and β-adrenoceptors［J］.Neurobiol Learn Mem,2023,205:107829.
［8］　ARIHARA Y,FUKUYAMA Y,KIDA S.Consolidation,reconsolidation,and extinction of contextual fear memory depend on de novo protein synthesis in the locus coeruleus［J］.Brain Res Bull,2023,202:110746.
［9］　GRFF J,JOSEPH N F,HORN M E,et al.Epigenetic priming of memory updating during reconsolidation to attenuate remote fear memories［J］.Cell,2014,156(1/2):261-276.
［10］　ENOMOTO S,KATO T A.Involvement of microglia in disturbed fear memory regulation:possible microglial contribution to the pathophysiology of posttraumatic stress disorder［J］.Neurochem Int,2021,142:104921.
［11］　LI S,LIAO Y,DONG Y,et al.Microglial deletion and inhibition alleviate behavior of post-traumatic stress disorder in mice［J］.J Neuroinflammation,2021,18(1):7.
［12］　PICARD K,CORSI G,DECOEUR F,et al.Microglial homeostasis disruption modulates non-rapid eye movement sleep duration and neuronal activity in adult female mice［J］.Brain Behav Immun,2023,107:153-164.
［13］　CUI X,ZHOU S,XIA G,et al.A multispecies probiotic accelerates fear extinction and inhibits relapse in mice:role of microglia［J］.Neuropharmacology,2021,193:108613.
［14］　PREZ-CABELLO J A,SILVERA-CARRASCO L,FRANCO J M,et al.MAPK/MAK/MRK overlapping kinase (MOK) controls microglial inflammatory/type-I IFN responses via Brd4 and is involved in ALS［J］.Proc Natl Acad Sci U S A,2023,120(28):e2302143120.
［15］　CHALLA N V D,CHEN S,YUAN H,et al.GSDMD gene knockout alleviates hyperoxia-induced hippocampal brain injury in neonatal mice［J］.J Neuroinflammation,2023,20(1):205.
［16］　LU X,ZHAN L,CHAI G,et al.Hypoxic preconditioning attenua-tes neuroinflammation via inhibiting NF-κB/NLRP3 axis mediated by p-MLKL after transient global cerebral ischemia［J］.Mol Neurobiol,2023,Epub ahead of print.
［17］　ZHANG W J,HUANG Y Q,FU A,et al.The retrotransposition of L1 is involved in the reconsolidation of contextual fear memory in mice［J］.CNS Neurol Disord Drug Targets,2021,20(3):273-284.
［18］　UWAYA A,LEE H,PARK J,et al.Acute immobilization stress following contextual fear conditioning reduces fear memory:timing is essential［J］.Behav Brain Funct,2016,12(1):8.
［19］　RAUT S B,MARATHE P A,VAN EIJK L,et al.Diverse therapeutic developments for post-traumatic stress disorder (PTSD) indicate common mechanisms of memory modulation［J］.Pharmacol Ther,2022,239:108195.
［20］　NADER K,SCHAFE G E,LE DOUX J E.Fear memories require protein synthesis in the amygdala for reconsolidation after retrieval［J］.Nature,2000,406(6797):722-726.
［21］　GRELLA S L,FORTIN A H,RUESCH E,et al.Reactivating hippocampal-mediated memories during reconsolidation to disrupt fear［J］.Nat Commun,2022,13(1):4733.
［22］　YU Z,FUKUSHIMA H,ONO C,et al.Microglial production of TNF-alpha is a key element of sustained fear memory［J］.Brain Behav Immun,2017,59:313-321.
［23］　MAURYA S K,GUPTA S,MISHRA R.Transcriptional and epigenetic regulation of microglia in maintenance of brain homeostasis and neurodegeneration［J］.Front Mol Neurosci,2023,15:1072046.
［24］　DEY A,YANG W,GEGONNE A,et al.BRD4 directs hematopoie-tic stem cell development and modulates macrophage inflammatory responses［J］.EMBO J,2019,38(7):e100293.
［25］　HOU J,HSU J M,HUNG M C.Molecular mechanisms and functions of pyroptosis in inflammation and antitumor immunity［J］.Mol Cell,2021,81(22):4579-4590.
［26］　PASPARAKIS M,VANDENABEELE P.Necroptosis and its role in inflammation［J］.Nature,2015,517(7534):311-320.
［27］　WANG J,CHEN J,JIN H,et al.BRD4 inhibition attenuates inflammatory response in microglia and facilitates recovery after spinal cord injury in rats［J］.J Cell Mol Med,2019,23(5):3214-3223.
［28］　GU L,SUN M,LI R,et al.Didymin suppresses microglia pyroptosis and neuroinflammation through the asc/caspase-1/GSDMD pathway following experimental intracerebral hemorrhage［J］.Front Immunol,2022,13:810582.
［29］　HE C,LIU Y,HUANG Z,et al.A specific RIP3+ subpopulation of microglia promotes retinopathy through a hypoxia-triggered necroptotic mechanism［J］.Proc Natl Acad Sci U S A,2021,118(11):e2023290118.
［30］　ZHOU Y,GU Y,LIU J.BRD4 suppression alleviates cerebral ischemia-induced brain injury by blocking glial activation via the inhibition of inflammatory response and pyroptosis［J］.Biochem Biophys Res Commun,2019,519(3):481-488.
［31］　XIONG Y,LI L,ZHANG L,et al.The bromodomain protein BRD4 positively regulates necroptosis via modulating MLKL expression［J］.Cell Death Differ,2019,26(10):1929-1941.
［32］　WANG Y,WERNERSBACH I,STREHLE J,et al.Early posttraumatic CSF1R inhibition via PLX3397 leads to time- and sex-dependent effects on inflammation and neuronal maintenance after traumatic brain injury in mice［J］.Brain Behav Immun,2022,106:49-66.
［33］　WYATT-JOHNSON S K,SOMMER A L,SHIM K Y,et al.Suppression of microgliosis with the colony-stimulating factor 1 receptor inhibitor PLX3397 does not attenuate memory defects during epileptogenesis in the rat［J］.Front Neurol,2021,12:651096.
［34］　ZHANG D,LI S,HOU L,et al.Microglial activation contributes to cognitive impairments in rotenone-induced mouse Parkinson′s disease model［J］.J Neuroinflammation,2021,18(1):4.
［35］　WANG L,WANG X,DENG L,et al.Pexidartinib (PLX3397) through restoring hippocampal synaptic plasticity ameliorates social isolation-induced mood disorders［J］.Int Immunopharmacol,2022,113(Pt B):109436.
［36］　DUAN Q,HUANG F L,LI S J,et al.BET proteins inhibitor JQ-1 impaired the extinction of remote auditory fear memory:an effect mediated by insulin like growth factor 2［J］.Neuropharmacology,2020,177:108255.
［37］　WANG W,WANG R,JIANG Z,et al.Inhibiting Brd4 alleviated PTSD-like behaviors and fear memory through regulating imme-diate early genes expression and neuroinflammation in rats［J］.J Neurochem,2021,158(4):912-927.
［38］　SHI J,ZHAO Y,WANG K,et al.Cleavage of GSDMD by infla-mmatory caspases determines pyroptotic cell death［J］.Nature,2015,526(7575):660-665.
［39］　MARTENS S,BRIDELANCE J,ROELANDT R,et al.MLKL in cancer:more than a necroptosis regulator［J］.Cell Death Differ,2021,28(6):1757-1772.
［40］　GERSTNER J R,YIN J C P.Circadian rhythms and memory formation［J］.Nat Rev Neurosci,2010,11(8):577-588.
［41］　KITAMURA T,OGAWA S K,ROY D S,et al.Engrams and circuits crucial for systems consolidation of a memory［J］.Science,2017,356(6333):73-78.
［42］　COLEMAN L G J,ZOU J,CREWS F T.Microglial depletion and repopulation in brain slice culture normalizes sensitized proinflammatory signaling［J］.J Neuroinflammation,2020,17(1):27.
［43］　SONG W,ZHU R,GAO W,et al.Blue light induces RPE cell necroptosis,which can be inhibited by minocycline［J］.Front Med,2022,9:831463.

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更新日期/Last Update: 2023-12-05