褪黑素对Alzheimer病模型大鼠认知功能和海马tau蛋白过度磷酸化的影响

目的 研究褪黑素(MT)对Alzheimer病(AD)模型大鼠认知功能和海马tau蛋白过度磷酸化的影响.方法 给大鼠海马内注射凝聚态β-淀粉样蛋白(Aβ)25-35制作AD模型;MT组大鼠从制模前7 d至制模后19 d每日腹腔注射MT,AD组大鼠制模后腹腔注射生理盐水;用Morris水迷宫试验检测大鼠的认知功能,银染法观察海马神经元形态,免疫组化法观察过度磷酸化tau蛋白的表达,并与正常对照组比较.结果 MT组大鼠Morris水迷宫试验结果明显好于AD组(均P＜0.001);海马CA1区磷酸化tau蛋白阳性细胞数(60.0±2.3)明显少于AD组(98.4±3.0)(P＜0.001),与正常对照组比较差异无统计学意义;海马CA1区神经元纤维形态较AD组规则.结论 MT可明显改善AD大鼠的认知功能,并且抑制海马tau蛋白的过度磷酸化.     

母婴分离应激对幼年大鼠脑衰反应调节蛋白2及微管动态性影响

目的探讨不同程度母婴分离(maternal separation,MS)应激对幼年雄性大鼠海马脑衰反应调节蛋白2(collapsin response mediator protein 2, CRMP2)及微管动态性的影响。方法 36只新生雄性大鼠随机分为3组,母婴分离360 min组(MS360组)、母婴分离15 min组(MS15组)及对照组(NC组)。出生后第4～10天进行1周的母婴分离,每天分离360 min或15 min,NC组正常饲养。采用q-RT-PCR技术检测海马CRMP2、actin、tubulin的mRNA表达水平,采用Western-Blot技术检测海马CRMP2、P-CRMP2、actin、tubulin及动态微管标志物Tyr-tubulin、稳定微管标志物Acet-tubulin表达水平。结果 3组海马P-CRMP2、Acet-tubulin、Tyr-tubulin表达水平差异有统计学意义(P0.05),MS360组与MS15组及NC组大鼠相比,P-CRMP2、Acet-tubulin表达水平升高(P0.05),Tyr-tubulin表达水平下降(P0.05),MS15组与NC组无统计学差异(P0.05)。3组CRMP2、actin、tubulin的mRNA和蛋白表达水平无统计学差异(P0.05)。结论长时间母婴分离应激可升高CRMP2的磷酸化水平,降低海马微管动态性,使神经可塑性受损,而短期母婴分离未对海马微管动态性产生不利影响。     

孕期酒精暴露对子代大鼠学习记忆及海马N-甲基-D-天冬氨酸受体2B亚基表达的影响

目的 研究孕期酒精暴露对子代大鼠学习记忆及海马N-甲基-D-天冬氨酸(NMDA)受体2B亚基(NR2B)表达的影响.方法 按照随机数字表法,将雌性SD大鼠随机分为正常对照组、饮酒对照组和孕期酒精暴露组,每组各8只;饮酒法建立大鼠孕期酒精暴露模型,子代成年后,采用Y-型迷宫测试子鼠学习记忆成绩;采用聚合酶链反应分析子鼠海马组织NR2B mRNA的表达;采用免疫荧光法检测子鼠海马区NR2B蛋白表达.结果 (1)各组子鼠成年后学习记忆成绩的差异有统计学意义(F=4.566,P＜0.05),孕期酒精暴露组子鼠学习记忆成绩[(43.00±15.33)次]比正常对照组[ (25.13±12.35)次]和饮酒对照组[(26.12±11.95)次]明显下降(P均＜0.05);(2)各组子鼠成年后海马组织中NR2B mRNA表达差异有统计学意义(F=29.795,P＜0.01),孕期酒精暴露组子鼠海马区NR2B mRNA表达(0.97±0.14)较正常对照组(0.52±0.10)和饮酒对照组(0.62±0.12)明显上升(P均＜0.01);孕期酒精暴露组子鼠海马区NR2B蛋白表达明显增加.结论 孕期酒精暴露对子代大鼠的神经损伤可能与NMDA受体亚基NR2B蛋白表达的上调有关.     

褪黑素对癫痫状态大鼠海马Caspase-3 mRNA表达和脂质过氧化的影响

目的 探讨褪黑素（melatonin,Mel）神经元保护作用的机制.方法 采用匹罗卡品（pilocarpine,PILO）诱导大鼠癫痫持续状态（status epilepticus,SE）模型,用比色法检测海马丙二醛（malondialdehyde,MDA）、还原型谷光甘肽（glutathione,GSH）水平和谷光甘肽还原酶（glutathione reductase,GR）的活力,用RT-PCR技术检测海马caspase-3 mRNA的表达.结果 PILO组大鼠SE后6h～72h,海马MDA含量明显高于对照组（P＜0.01）;海马GSH含量和GR活力均明显低于对照组（P＜0.01）,SE后7d,海马MDA含量和GR活力基本恢复正常.PILO＋Mel组大鼠,在SE后各时相点海马MDA含量均明显低于PILO组大鼠（P＜0.01）;而海马GSH含量和GR活力均显著高于PILO组大鼠（P＜0.05）.SE后6h～72h,PILO组大鼠海马caspase-3 mRNA的表达明显高于对照组（P＜0.01）和PILO＋Mel组（P＜0.01）,提示给予Mel可明显抑制SE大鼠海马caspase-3 mRNA的表达.结论 Mel发挥神经元保护作用的机制是通过提高SE大鼠海马GSH水平和GR活力,来减轻脂质过氧化损伤;并通过抑制caspase-3 mRNA的表达,来干预细胞凋亡.     

捕食应激致大鼠持续性情绪唤醒障碍时血浆皮质酮及海马皮质类固醇受体表达变化

目的 探讨严重心理应激所致情感行为异常的相关神经生物学机制.方法 将136只雄性Wistar大鼠随机分为捕食应激组(以下简称应激组)和正常对照组(以下简称对照组),每组68只.以放射免疫分析法检测应激后1 h、1 d、1周,以及1,2,4个月时血浆皮质酮水平,并采用免疫组化和蛋白质免疫印迹法,检测海马糖皮质激素受体(GR)及盐皮质激素受体(MR)表达的变化规律.结果 (1)血浆皮质酮水平:应激后1 h应激组[(44.5±9.3)μg/L]高于对照组[(22.6±4.0)μg/L,P＜0.01],但应激后1 d降至正常水平,而应激后1周至4个月则明显低于对照组[应激组分别为(μg/L)12.4±2.5,9.8±2.1,8.7±2.1,10.1±2.3,对照组分别为(μg/L)20.8±3.9,21.1±4.2,16.6±3.6及20.2±4.0;P＜0.01].(2)海马GR和MR阳性免疫反应信号:应激后1 d免疫组化研究显示,应激大鼠海马及额叶皮层GR均较对照组增强(P＜0.01),其中以海马结构的改变更明显(P＜0.05);而MR则显著降低(P＜0.01).海马免疫印迹检测进一步揭示,应激组海马GR表达于应激后1 d至4个月明显增高(P＜0.05),而MR表达则于应激后1 h至1个月显著低于对照组(P＜0.05).结论 捕食应激后大鼠出现血浆皮质酮双向变化、海马GR和MR失衡以及不同的变化规律,这种严重的心理应激在引发大鼠持续性情绪唤醒障碍中可能有重要作用.     

创伤性脑损伤后大鼠海马区糖皮质激素受体mRNA的表达

目的研究创伤性脑损伤(TBI)后大鼠海马区糖皮质激素受体(GR)mRNA表达的变化及其对大鼠认知功能的影响。方法建立大鼠头颅侧向旋转加速脑创伤模型,应用逆转录酶-聚合酶链式反应(RT-PCR)和Morris水迷宫检测伤后大鼠海马区GR mRNA的表达与学习记忆功能的关系。结果伤后4～7 d大鼠海马区GR持续低表达;Morris水迷宫检测伤后大鼠出现认知功能障碍。结论 TBI大鼠海马区GR mRNA的降低影响大鼠认知功能。     

多发性硬化患者糖皮质激素受体蛋白和mRNA表达与激素疗效的相关性

目的 检测多发性硬化(MS)患者外周血单个核细胞(PBMC)中糖皮质激素受体(GR)蛋白与mRNA亚型:GRα mRNA、GRβ mRNA的表达,探讨GR蛋白和GRα mRNA、GRβ mRNA的表达与甲泼尼龙静脉冲击治疗(IVMP)疗效间的关系.方法 采用放射配体结合法测定20例缓解复发型MS(RRMS)、6例继发进展型MS(SPMS)及26名健康对照GR蛋白数量;采用半定量逆转录聚合酶链反应法测定GRα mRNA、GRβ mRNA的表达,使用扩展残疾状态量表(EDSS)评价IVMP疗效.结果 (1)IVMP前RRMS和SPMS患者GR蛋白数量分别为(3.8±0.2)×103位点/细胞和(1.6±0.2)×103位点/细胞,均显著低于健康对照组[(4.2±0.8)×103位点/细胞,P＜0.05].RRMS患者IVMP前GR蛋白数量与EDSS呈线性负相关(r=-0.441,P=0.015).(2)以磷酸甘油醛脱氢酶的表达作为内参照,RRMS患者GRα mRNA的表达水平(0.792±0.177)与健康对照组(0.805±0.158)差异无统计学意义,而SPMS患者的表达水平(0.315±0.129)明显低于健康对照(P＜0.05).(3)RRMS和SPMS患者PBMC中均有GRβ mRNA的表达,健康对照组未检出GRβ mRNA表达.(4)RRMS、SPMS患者GRβ mRNA的表达分别占各自GRα mRNA的43.98%±2.40%和140.01%±78.75%.结论 RRMS患者GRβ mRNA的低水平表达不影响IVMP疗效.SPMS患者GRα mRNA表达水平显著降低和(或)GRβ mRNA表达水平显著升高导致GC抵抗.     

轻型颅脑损伤对大鼠脑源性神经营养因子基因表达的影响及意义

目的探讨轻型颅脑损伤对大鼠脑源性神经营养因子(BDNF)基因表达的影响及意义.方法120只成年SD大鼠根据性别不同分为雌性对照组、雌性实验组、雄性对照组和雄性实验组,每组30只.两实验组建立侧位液压冲击轻型颅脑损伤模型.Morris水迷宫试验测试大鼠学习记忆能力,RT-PCR检测海马中BDNF mRNA的表达.结果水迷宫定位航行试验第4天,同性别大鼠实验组平均逃逸潜伏期较对照组明显延长(P＜0.05);空间探索试验同性别大鼠实验组寻找平台潜伏期较对照组明显延长(P＜0.05),穿越平台次数较对照组减少(P＜0.05);同性别大鼠实验组大脑海马中BDNFmRNA的表达显著低于对照组(P＜0.01).结论侧位液压冲击轻型颅脑损伤会降低模型鼠大脑海马中BDNF mRNA的表达,并可能因此导致大鼠学习记忆能力下降.     

创伤后应激障碍大鼠海马组织5-HT1受体mRNA的表达

目的探讨创伤后应激障碍(Posttraumatic Stress Disorder,PTSD)大鼠海马组织中5-羟色胺受体1mRNA的表达。方法随机将雄性wistar大鼠分为对照组和PTSD组,采用SPS(single-prolonged stress)方法刺激大鼠建立PTSD大鼠模型,Morris水迷宫实验检测大鼠记忆功能,化学发光法检测大鼠血清皮质醇浓度。SPS刺激后1d、4d、7d、14d分离海马组织,RT-PCR方法检测5-羟色胺受体1的变化。结果 (1)对照组水迷宫实验逃避潜伏期为(5.632±1.065)s,模型组为(20.762±3.236)s(t=9.932,P0.01);(2)对照组血清皮质醇浓度为(1.25±0.12)μg/dl,模型组为(0.58±0.09)μg/dl(t=7.340,P0.01);(3)SPS刺激后1d、4d、7d、14d海马5-HT1A受体mRNA表达相对水平分别为0.846±0.067、0.510±0.052、1.007±0.137、1.109±0.106,与对照组相1.223±0.152比较,1d、4d、7d表达低于对照组(P0.05);5-羟色胺1受体其他亚型未见表达。结论 PTSD大鼠海马中5-HT1A受体mRNA表达降低。     

吗啡戒断性焦虑大鼠海马突触界面结构及突触素表达变化

目的 探讨吗啡依赖戒断焦虑行为与海马CA1、CA3区突触界面结构和突触素表达变化之间的相关性.方法 剂量递增法建立大鼠吗啡依赖模型,高架十字迷宫检测焦虑行为,透射电镜技术结合图像分析系统、免疫组织化学比较对照组、模型组和治疗组(各6只)大鼠海马CA1、CA3区突触界面结构和突触素(P38)的表达.结果 (1)行为学:模型组开放臂的次数和时间均少于对照组和治疗组[最小有意义差异t检验(下同),P＜0.01或P＜0.05).(2)突触界面结构:模型组CA1区突触后致密物厚度[(10.7±0.9)nm]、突触活性区长度[(45±4)am]、突触间隙宽度[(3.80±0.30)nm]和突触界面曲率(1.37±0.12)均高于对照组和治疗组(P＜0.01或P＜0.05);模型组CA3区突触后致密物厚度[(12.7±1.1)nm]、突触活性区长度[(53±8)nm]、突触间隙宽度[(3.81 ±0.59)nm]、突触界面曲率(1.39±0.30)亦均高于对照组和治疗组(P＜0.01或P＜0.05).(3)突触素表达:模型组CA1、CA3区突触素吸光度(A)值分别为(0.42±0.06)和(0.43±0.05),显著高于对照组(0.2±0.02,0.25±0.03)和治疗组(0.27±0.04,0.26±0.03).结论吗啡戒断焦虑行为与海马CA1、CA3区突触形态结构可塑性及突触素表达水平有一定的相关性.     

Periodic maternal separation decreases hippocampal neurogenesis without affecting basal corticosterone during the stress hyporesponsive period, but alters HPA axis and coping behavior in adulthood

Although not directly evaluated, the early rise of glucocorticoid (GC) levels, as occur after exposure to adverse early life experience, are assumed to affect hippocampal ontogeny by altering the hippocampus negative feedback on adult HPA axis. To test whether hippocampal ontogeny is affected by early exposure to stress we estimated the survival of recently formed hippocampal granule cells in rat pups subjected to periodic maternal separation (180 min/day; MS180) from postnatal days (PND) 1 to 14. Accordingly, MS180 pups injected with bromodeoxyuridine (BrdU, 50 mg/kg, ip) at PND 5 showed decreased density of doublecortin (DCX) positive BrdU-labeled cells at PND 15. MS180 and AFR pups showed similar corticosterone (CORT) basal levels between PND 3 and 12, whereas adult MS180 rats presented with higher CORT levels than AFR adults. Nonetheless, both AFR and MS180 pups and adults showed similar transient increments of CORT levels in response to stress. In addition, MS180 had no effect on the adult anxiety-like behavior evaluated in the elevated plus maze, but evoked a passive coping strategy in the forced swimming test. The data show that the decrease in hippocampal neurogenesis is an early onset phenomenon, and suggests that adverse experiences alter hippocampal ontogeny without chronic elevation of GC levels.     

Neonatal maternal separation alters the development of glucocorticoid receptor expression in the interpositus nucleus of the cerebellum

Adverse early experience impairs adult learning and memory. Previously, we showed that neonatal maternal separation impaired eyeblink conditioning in adult male rats. This impairment was correlated with increases in glucocorticoid receptor expression in the posterior region of the cerebellar interpositus nucleus, a key structure in the neural circuitry controlling eyeblink conditioning. To begin to establish how separation results in altered glucocorticoid receptor expression in adulthood, we assessed the developmental pattern of glucocorticoid receptor expression in the interpositus nucleus in controls versus rats that had undergone maternal separation for 1 h per day on postnatal days 2–14. Rat pups were exposed to either standard rearing (control) or maternal separation and glucocorticoid receptor expression was assessed at postnatal day 15, postnatal day 21, and adulthood. In control males, glucocorticoid receptor expression in the interpositus nucleus declined between postnatal days 15 and 21, then increased into adulthood. On postnatal day 15, there was less glucocorticoid receptor expression in the interpositus nucleus in males that were maternally separated than in controls. However, neonatal separation significantly attenuated the normal decline in the third postnatal week, resulting in significantly greater glucocorticoid receptor expression in the interpositus in separated males than in control rats at postnatal day 21. The developmental pattern of glucocorticoid receptor expression was not altered by maternal separation in female rats. Thus, maternal separation may impair learning and memory in adult males by altering normal developmental changes in glucocorticoid receptor expression.     

Pinealectomy stimulates and exogenous melatonin inhibits harmful effects of epileptiform activity during pregnancy in the hippocampus of newborn rats: an immunohistochemical study

Objectives Epilepsy during the pregnancy is an important problem in clinical practice for newborn individuals. Recently, it has been demonstrated that mothers’ epileptic seizures have some harmful effects on newborns, but present data concerning the effects of epileptic phenomena in pregnant mothers on newborn pups are still limited. The current study was undertaken to investigate the morphological changes in the hippocampus of newborn pups of pinealectomized rats subjected to experimental epilepsy during pregnancy.Methods In this study, rats were randomly divided into four groups (ten animals each): intact control group, epilepsy control group, surgical pinealectomy + epilepsy group, and group with melatonin treatment following pinealectomy procedure. The animals in surgical pinealectomy + epilepsy and melatonin treatment groups underwent a surgical intervention consisting of pineal gland removal. At 1 month after surgical pinealectomy, an acute grand mal epileptic seizure was induced by 400 IU penicillin G administration into their hippocampal CA3 region on the 13th day of their pregnancy in all animals except the intact control animals. On the first neonatal day, the hippocampi were removed and processed for microscopic examination. Nestin expression was analysed in the developing hippocampal tissue.Results Normal migration and hippocampal maturation were determined in the postnatal rat hippocampus in intact control group, but the morphological structure of the hippocampus in the epilepsy control group corresponded to the early embryonal period. It was found that experimental epilepsy and pinealectomy enhanced nestin immunoreactivity, whereas exogenous melatonin treatment (30 μg/100 g body weight, intraperitoneal) inhibited pinealectomy-stimulated nestin expression in CA1 region of the hippocampus.Conclusion These findings suggest that epileptic seizures during pregnancy may cause an impaired hippocampal neurogenesis and neuronal maturation in the newborn, and the negative effects in the postnatal rat hippocampus are more dramatic after pinealectomy of the mother; conversely, melatonin administration suppresses these negative changes. This is the first report investigating the effects of maternal epilepsy during pregnancy in pinealectomized rats on nestin immunoexpression in the newborn rat hippocampus.Presented in part at the 4th Asian-Pacific International Congress of Anatomists (APICA), Kuşadası, Turkey, 7-10 September 2005.     

Cognitive impairment associated to HPA axis hyperactivity after maternal separation in rats

Exposure to early stressful adverse life events may increase vulnerability to psychopathology in adult life. There are important memory disturbances in stress-related psychiatric disorders. Therefore, there is much interest in understanding the mechanisms responsible for interactions between stress and cognition. Male Wistar rats that experienced 3-h daily separations from the dam during the first 3 weeks of life (maternal separation, MS) showed in adulthood a depressive-like behaviour in the forced swimming test, increased hypothalamic-pituitary-adrenal (HPA) axis responsiveness to stressors and elevated CRF mRNA in the paraventricular nucleus of the hypothalamus (PVN). In the hippocampus of MS rats, there was a lower glucocorticoid receptor density. MS produced significant learning impairments both in the Morris water maze and in the novel object recognition test (NORT). The glucocorticoid receptor antagonist mifepristone and the beta-adrenoceptor antagonist propranolol were able to completely reverse the increased immobility time in the forced swimming test and the memory deficits in the NORT observed in MS rats. Our data support the hypothesis that elevated secretion of glucocorticoids may be associated to behavioural and cognitive deficits in MS rats. The stress hyperresponsiveness observed in MS rats could be attributed, at least in part, to an impaired feedback sensitivity mediated by hippocampal glucocorticoid receptors. It can also be suggested the possible involvement of the noradrenergic system in cognitive impairments mediated by glucocorticoids in the MS model.     

重复经颅磁刺激对抑郁模型大鼠行为学及海马区糖皮质激素受体表达的影响

目的观察重复经颅磁刺激(repetitive transcranial magnetic stimulation,r TMS)对慢性应激抑郁模型大鼠的抗抑郁作用及对海马区糖皮质激素受体(glucocorticoid receptor,GR)表达的影响,探讨r TMS抗抑郁作用的可能机制。方法 75只健康成年雄性大鼠随机分为造模组(60只)和空白对照组(15只),造模组采用孤养联合慢性温和不可预见应激(chronic unpredictability stimulus,CUMS)方法制备抑郁大鼠模型,为期3周,筛选造模成功的大鼠45只随机分为r TMS组、伪r TMS组和抑郁对照组,每组15只,r TMS组和伪r TMS组分别接受10 Hz的r TMS刺激和伪刺激干预3周,抑郁对照组和空白对照组不给予干预。分别于造模前、造模后、r TMS干预后进行体重测量、蔗糖水消耗实验和强迫游泳实验评估,r TMS干预后检测大鼠海马区GR蛋白和海马GR m RNA表达水平。结果造模后,r TMS组、伪r TMS组和抑郁对照组大鼠蔗糖水消耗量较空白对照组下降,强迫游泳不动时间增加(P0.01)。r TMS干预后,r TMS组体重增长率、蔗糖水消耗量与伪r TMS组和抑郁对照组相比均较高(P0.01),强迫游泳不动时间较短(P0.01)。伪r TMS组及抑郁对照组海马区GR蛋白及其m RNA表达水平与r TMS组和空白对照组相比均较低(P0.05)。结论 r TMS能够改善CUMS抑郁模型大鼠的抑郁样行为,可能与上调海马区GR表达有关。     

Distribution of glucocorticoid and mineralocorticoid receptor messenger RNA expression in human postmortem hippocampus

Corticosteroids bind to hippocampal glucocorticoid (GR) and mineralocorticoid (MR) receptors, thereby affecting behaviour and neurochemical transmission. Rat hippocampus has high levels of both receptors and their messenger RNAs (mRNA), but there is little information on receptors in human brain. We used in situ hybridization to determine the distribution of GR and MR mRNA expression in human hippocampus. Frozen sections of human postmortem hippocampus (5 patients, 58-88 years old, without cerebral pathology) were postfixed in paraformaldehyde and hybridized with 35S-UTP-labelled cRNA probes (transcribed in vitro from human cDNA subclones) under stringent conditions. Control included hybridization with sense probes and heterologous cRNA competition studies. GR mRNA was highly expressed in dentate gyrus, CA3 and CA4, but levels were significantly lower in CA1 and CA2. MR mRNA was also very highly expressed in hippocampus, with significantly higher levels in dentate gyrus and CA2, CA3 and CA4 than CA1. Controls confirmed the specificity of hybridization and there was little hybridization of sense probes. High GR and MR mRNA expression is found in both rat and human hippocampus but the subregional distributions clearly differ between the species.     

Morphine exposure prevents up-regulation of MR and GR binding sites in the brain of adult male and female rats due to prenatal stress

Our previous work demonstrated that the hormone response to stress and the negative feedback inhibition to these hormones are sex-dependently altered by prenatal morphine exposure in adult rats. An alteration in the glucocorticoid negative feedback inhibition is mediated by glucocorticoid receptors (GR) that are distributed throughout the brain, and mineralocorticoid receptors (MR) localized mainly in the hippocampus and involved in a tonic influence of brain functions. Therefore, the present study examined the binding characteristics of MR and GR in young adult male and female rats exposed prenatally (E11-E18) to morphine (10 mg/kg/2 x /day), saline or no treatment at all (controls). At 60-90 days of age, animals were adrenalectomized (ADX) 24 h prior to decapitation. The hippocampus and hypothalamus were dissected for saturation binding assays. The data demonstrate that prenatal stress due to maternal saline injections up-regulates MR and GR binding in the hippocampus of adult male rats and this effect is prevented by prenatal morphine exposure. There is no effect of prenatal morphine exposure on GR binding in the hypothalamus of males. In female rats, prenatal morphine exposure does not affect the binding of MR and GR in the hippocampus or GR in the hypothalamus relative to controls; however, they are affected by ovarian hormone fluctuation. Moreover, prenatal stress decreases MR binding in the hippocampus of diestrous females and GR binding in the hypothalamus of estrous females. Both decreases are prevented by prenatal morphine exposure. Thus, the present study demonstrates that: (1) prenatal stress due to maternal saline injections alters MR and GR binding of adult male and female rats and is prevented by prenatal morphine exposure; (2) the MR and GR binding in adult female rats are affected by ovarian hormone fluctuations.     

Neonatal maternal separation reduces hippocampal mossy fiber density in adult Long Evans rats

Neonatal maternal separation of rat pups leads to a stable stress hyper-responsive phenotype characterized by increased basal levels of corticotropin releasing factor (CRF) mRNA in the hypothalamic and extra-hypothalamic nuclei, increased hypothalamic CRF release, and enhanced adrenocorticotrophin hormone (ACTH) and corticosterone (CORT) responses to psychological stressors. Stress and exposure to glucocorticoids either early in life or in adulthood have been associated with hippocampal atrophy and impairments in learning and memory. In this study, male Long Evans rat pups were exposed to daily 3-h (HMS180) or 15-min (HMS15) periods of maternal separation on postnatal days (PND) 2–14 or normal animal facility rearing. Maternal separation and subsequent reunion with the dam resulted in elevated plasma CORT levels versus HMS15 animals at PND7, a time when rat pups are normally hyporesponsive to stressors and show limited pituitary–adrenal responses. As adults, HMS180 rats exhibited elevated indices of anxiety, startle-induced pituitary–adrenal hyper-responsiveness, and slight, but significant impairment on acquisition in the Morris water maze task. In addition, HMS180 rats exhibited decreased mossy fiber density in the stratum oriens region of the hippocampus as measured by Timm’s staining, but no change in volume of the dentate gyrus. These changes may be the result of neonatal exposure to elevated glucocorticoids and/or changes in other signaling systems in response to maternal separation. Overall the results suggest that repeated, daily, 3-h maternal separations during critical periods of hippocampal development can disrupt hippocampal cytoarchitecture in a stable manner. The resulting change in morphology may contribute to the subtle, but consistent learning deficit and overall stress hyper-responsive phenotype observed in these animals.     

Maternal separation activates microglial cells and induces an inflammatory response in the hippocampus of male rat pups,independently of hypothalamic and peripheral cytokine levels

Adult animals subjected to chronic stress show an inflammatory response in the hippocampus which has been related to cognitive dysfunction and psychopathology. However the immediate consequences of early life stress on hippocampal glial cells have not been studied. Here we analyzed the effects of maternal separation (MS) on astrocyte and microglial cell morphology in the hippocampal hilus, compared the expression of cytokines in the hippocampus and hypothalamus, and the peripheral response of cytokines, on postnatal day (PD) 15.Male rat pups of MS (3 h/day, PD1–PD14) and Control (CONT) pups showed similar microglial cell densities in the hilus, but MS pups presented more activated microglia. MS decreased astrocyte density and the number of processes in the hilus. Cytokine mRNA expression (qPCR) was analyzed in MS and CONT groups, sacrificed (i) under basal (B) conditions or (ii) after a single stress event (SS) at PN15. In hippocampal extracts, MS increased IL-1β mRNA, under B and SS conditions while IL-6 and TNF-α did not change. In hypothalamic tissue, MS increased TNF-α and IL-6 mRNA, but not IL-1b, after SS. Peripheral concentrations of IL-1β were decreased under B and SS conditions in MS; IL-6 concentration increased after SS in MS pups, and TNF-α concentration was unchanged. In conclusion, MS activates microglial cells and decreases astrocyte density in the hippocampus. A differential cytokine expression is observed in the hippocampus and the hypothalamus after MS, and after SS. Also, MS triggers an independent response of peripheral cytokines. These specific responses together could contribute to decrease hippocampal neurogenesis and alter the neuroendocrine axis.