解郁丸对WKY抑郁样大鼠前额叶皮层与海马树突棘的影响

目的:观察Wistar-Kyoto(WKY)抑郁模型大鼠前额叶皮层与海马组织中树突棘的变化,并探讨解郁丸在其中的作用。方法:以成年雄性WKY大鼠为实验组,选用同品系SPF级Wistar大鼠为其对照组。首先,用糖水偏好实验、旷场实验和强迫游泳实验检测大鼠行为学变化,并作为其基线;然后将实验组所有WKY大鼠随机分为模型组、解郁丸组和西酞普兰组,与对照组分别进行21 d灌胃给药,再用相同的行为学方法检测给药后行为的变化。采用Golgi染色观察前额叶皮层与海马组织中树突棘的病理特点。Western blot检测前额叶皮层与海马组织突触后致密区蛋白95(PSD-95)的表达水平。结果:给药前,WKY大鼠表现出明显的抑郁样行为,前额叶皮层与海马组织中树突棘的密度明显降低(P<0. 01),且PSD-95蛋白的表达水平明显降低(P<0. 01);经药物治疗后,WKY大鼠的抑郁样行为明显减少,前额叶皮层与海马组织中树突棘的密度升高(P<0. 01),PSD-95蛋白的表达水平也提高(P<0. 01)。结论:解郁丸能明显减少WKY大鼠抑郁样行为,且影响树突棘的结构改变及PSD-9...     

MyD88抑制肽对小鼠脑缺血后小胶质细胞极化的影响

目的:研究髓样分化因子88抑制肽(MIP)对小鼠脑缺血后小胶质细胞极化的影响。方法:将30只雄性C57BL/6小鼠分为假手术组(sham)、局灶性脑缺血组(FCI)和给药组(MIP)。用光化学法建立小鼠FCI模型,给药组于造模后第3 d开始给予腹腔注射MIP,每次10 mg/kg,每日一次,连续3 d。采用real time RT-PCR检测损伤区白细胞介素(IL)-1β、IL-4、IL-10、肿瘤坏死因子α(TNF-α)、CD86和CD206 mRNA的表达,用Western Blot检测损伤区局部Toll样受体2(TLR2)、TLR4、髓样分化因子88(MyD88)、诱导型一氧化氮合酶(i NOS)和精氨酸酶-1(Arg-1)蛋白的表达水平,用免疫荧光染色观察损伤区i NOS和Arg-1与F4/80或Iba-1的共标情况。结果:与假手术组相比,FCI组小鼠损伤区MyD88、TLR2和TLR4蛋白的表达水平明显升高。与FCI组相比,MIP给药组小鼠损伤区IL-1β、TNF-ɑ和CD86 mRNA,以及iNOS蛋白的表达量明显下降,而IL-4、IL-10和CD206 mRNA,以及Ar...     

腹内侧前额叶皮质锥体神经元参与选择性坐骨神经损伤小鼠疼痛和焦虑样行为调控的研究

目的 探究腹内侧前额叶皮质（ventromedial prefrontal cortex,vmPFC）锥体神经元对选择性坐骨神经损伤（spared nerve injury,SNI）小鼠疼痛和焦虑样行为的调控作用。方法 构建SNI神经病理性疼痛（neuropathic pain,NP）动物模型,假手术组（sham）仅暴露神经。向SNI小鼠损伤对侧腹内侧前额叶皮质（ventromedial prefrontal cortex,vmPFC）注射红藻氨酸/生理盐水,或化学遗传学抑制病毒/空载对照病毒。采用von frey纤维丝检测SNI术后损伤侧机械缩足反射阈值（paw withdrawl threshold,PWT）;高架十字迷宫评估SNI术后焦虑情况;免疫荧光染色观察SNI损伤对侧vmPFC中c-fos表达及注射红藻氨酸后神经元毁损、星形胶质细胞活化情况,RNA-scope原位杂交染色与免疫荧光染色双标明确Vgat基因和Vglut1基因分别与c-fos的共表达情况。结果 术后7 d及14 d,相比于sham小鼠,SNI小鼠损伤侧足底PWT、进入开臂次数和停留时间明显减少（P<0.0...     

小胶质细胞M2型极化相关的调节分子研究进展

<正>中枢神经系统(central nervous system,CNS)的固有免疫反应是对损伤、炎症以及退行性疾病产生的首要免疫应答。多种CNS疾病,如创伤、脑炎、帕金森病(Parkinson's disease,PD)、阿尔茨海默症(Alzheimer's disease,AD)、多发性硬化(multiple sclerosis,MS)等,其发展转归与CNS的免疫与炎症变化有着密切联系。由于免疫与炎症反应的双刃剑效应,对CNS的免疫炎症的调控一直难以解决,而近年来对CNS定居的非专职免疫细胞-小胶质细胞的极化现象研究为CNS固有免疫反应的调节带来新的解题思路。     

槲皮素对小鼠前额叶皮质锥体神经元的放电抑制作用研究

目的:探讨槲皮素对小鼠前额叶皮质(prefrontal cortex,PFC)锥体神经元动作电位的效应。方法:成年C57小鼠或Thy1-GCaMP 3.0小鼠,制作含PFC的冠状脑片。分成正常脑脊液孵育组(对照组)和槲皮素孵育组(50μmol/L),给予高钾(5 mmoL/L KCl)人工脑脊液灌流两组脑片记录GCaMP荧光信号,全细胞膜片钳记录两组PFC锥体神经元放电,碘化丙啶(propidium iodide,PI)染色检测神经元活性。结果:5 mmol/L KCl人工脑脊液可以明显引起对照组GCaMP脑片PFC锥体神经元荧光信号增强,而对槲皮素组无明显作用。全细胞膜片钳记录显示槲皮素组神经元能够产生动作电位。但与对照组相比,产生放电所需刺激强度增加,连续放电能力降低。PI染色显示槲皮素组神经元死亡数目减少。结论:槲皮素孵育可以通过抑制小鼠前额叶皮质锥体神经元产生动作电位的能力,从而提高神经元在体外环境的生存能力。其具体的离子通道机制及细胞信号通路研究需进一步的深入研究。     

依达拉奉对脂多糖激活的小胶质细胞向M2型极化的影响

目的:探究依达拉奉对脂多糖(LPS)激活的小胶质细胞(MG)向M2型极化的影响。方法:利用脂多糖(LPS)激活体外培养的BV2小胶质细胞,复制小胶质细胞激活模型,分为对照组(control)、模型组(LPS)和LPS+依达拉奉组(LPS+E,100μmol/L),通过Western Blot和荧光双标染色技术,检测M2型小胶质细胞标记物：几丁质酶样蛋白1/2 (YM1/2)、精氨酸酶-1 (Arg-1)、白介素10 (IL-10)和甘露糖受体(CD206)的表达变化。结果:Western Blot和免疫荧光双标染色均提示：LPS组中YM1/2、Arg-1、IL-10和CD206的表达升高,与control组相比差异具有显著性(P<0.05),依达拉奉干预后,M2型小胶质细胞标记物表达进一步增高,LPS+E组与LPS组相比差异具有统计学意义(P<0.05)。结论:依达拉奉可促进LPS激活的小胶质细胞向M2型极化,发挥抗炎作用。     

延髓小胶质细胞极化参与实验性牙髓炎疼痛的研究

目的:检测小鼠实验性牙髓炎发生、发展过程中,延髓背角小胶质细胞的极化状态及其变化,探究延髓背角小胶质细胞不同极化状态与牙髓炎疼痛的相关性。方法:随机抽样法将C57小鼠分为对照组(Control)和实验性牙髓炎症模型组(Pulpitis)。牙髓暴露法建立实验性牙髓炎症模型,HE染色观察不同时间点牙髓炎症的变化情况;擦面实验观察小鼠牙髓炎性痛程度;Western Blot检测小胶质细胞特异性标志物离子钙结合衔接分子1(Iba-1)、M1型小胶质细胞极化标记物诱导型一氧化氮合酶(iNOS)及M2型小胶质细胞极化标记物精氨酸酶1(Arg-1)在延髓背角的表达及变化;免疫荧光组织化学染色法观察Iba-1、iNOS、Arg-1的表达及定位情况;real time RT-PCR检测IL-1β和IL-4 mRNA的表达变化。结果:HE染色结果显示,牙髓暴露1 d小鼠牙髓组织出现明显的炎性反应,3 d炎性发展至整个冠髓,7 d炎症扩散至根髓,14 d牙髓几乎全部坏死。痛行为学检测结果显示,牙髓暴露后1 d疼痛最为明显,随后开始逐渐减弱,7 d后恢复至正常水平。Western Blot、免疫荧光组织化学染...     

缺血性脑损伤状态下小胶质细胞的极化现象与影响因素

<正>小胶质细胞(microglia,MG)是中枢神经系统(central nervous system,CNS)的一类主要的胶质细胞,约占所有胶质细胞的5%~20%[1]。MG在脑中分布广泛,有明显的种属差异,并且在同一种属内也有区域差异,人脑中白质多于灰质,以延髓和脑桥密度最高,小脑灰质密度最低,较低的还有额叶、顶叶和枕叶~([1,2]);成年啮齿类动物脑内,灰质多于白质,以海马、嗅觉端脑、基底神经节和黑质的密度最高,纤维束、     

吲哚丙酸抑制小胶质细胞M1极化治疗脊髓损伤

背景：吲哚丙酸被证明可减轻糖尿病所致的中枢神经系统炎症,但其能否抑制小胶质细胞M1极化治疗脊髓损伤,目前仍缺乏相关研究。目的：通过细胞实验及动物实验探究吲哚丙酸抑制小胶质细胞M1极化治疗脊髓损伤的机制。方法：(1)体外实验：CCK8法检测BV2细胞活性并筛选最佳的吲哚丙酸使用浓度;然后将BV2细胞分为对照组、单纯给药组(50μmol/L吲哚丙酸)、脂多糖组(100 ng/mL脂多糖)、治疗组(100 ng/mL脂多糖+50μmol/L吲哚丙酸),采用Griess法检测一氧化氮含量;实时荧光定量PCR、Western Blot检测促炎相关因子的mRNA和蛋白的表达;细胞免疫荧光染色检测诱导型一氧化氮合酶的表达;Seahorse实验检测BV2细胞糖酵解压力水平。(2)体内实验：将30只SD大鼠随机分成3组：假手术组、脊髓损伤组、吲哚丙酸组。采用BBB评分与斜板实验评估大鼠脊髓损伤后功能恢复情况;免疫荧光染色检测脊髓组织中小胶质细胞诱导型一氧化氮合酶的表达情况;ELISA检测脊髓组织中促炎因子白细胞介素1β及肿瘤坏死因子α蛋白表达水平。结果与结论：(1)体外实验：当吲哚丙酸浓度> 50...     

Chronic stress effects on dendritic morphology in medial prefrontal cortex: sex differences and estrogen dependence

A growing body of work has documented sex differences in many behavioral, neurochemical, and morphological responses to stress. Chronic stress alters morphology of dendrites in medial prefrontal cortex in male rats. However, potential sex differences in stress-induced morphological changes in medial prefrontal cortex have not been examined. Thus, in Experiment 1 we assessed dendritic morphology in medial prefrontal cortex in male and female rats after chronic stress. Male and female rats underwent either 3 hours of restraint daily for 1 week or were left unhandled except for weighing. On the final day of restraint, all rats were euthanized and brains were stained using a Golgi–Cox procedure. Pyramidal neurons in layer II–III of medial prefrontal cortex were drawn in three dimensions, and morphology of apical and basilar arbors was quantified. In males, stress decreased apical dendritic branch number and length, whereas in females, stress increased apical dendritic length. In Experiment 2, we assessed whether estradiol mediates this stress-induced dendritic hypertrophy in females by assessing the effects of restraint stress on female rats that had received either ovariectomy with or without 17-β-estradiol replacement or sham ovariectomy. Brains were processed and neurons reconstructed as described in Experiment 1. Both sham-operated and ovariectomized rats with estradiol implants showed stress-induced increases in apical dendritic material, whereas ovariectomy without estradiol replacement prevented the stress-induced increase. Thus, the stress-induced increase in apical dendritic material in females is estradiol-dependent.     

Selective neonatal depletion of dopamine has no effect on medial prefrontal cortex self-stimulation in the rat

The role of the dopaminergic input to the medial prefrontal cortex (MFC) on self-stimulation (SS) was investigated in adult rats injected neonatally with 6-hydroxydopamine (6-OHDA). Each subject on day 3 or 5 received bilateral intraventricular injection of 6-OHDA (total dose 200 micrograms, 50 micrograms/injection/2.5 microliters vehicle which contained 1 mg/ml ascorbic acid) or of the vehicle alone after pretreatment with desmethylimipramine (50 mg/kg i.p.) 30 min earlier. At 150 days of age, the animals were implanted with monopolar (100 microns) stainless steel electrodes in the MFC. One long (10 h) and 5 short (2 h) SS sessions resulted in similar percentages of responders for the brain reward in test and control subjects, and similar response rates in both groups. Biochemical assays of the levels of norepinephrine (NE) and dopamine (DA) in the frontal cortex showed depletion of DA 90% in the test animals, but no depletion of NE. Histochemical fluorescence visualization of the catecholamine input verified the biochemical results in the MFC. These results are viewed as negative evidence for the hypothesis that DA innervations in the MFC are critical neural substrates for SS, and suggest that activation of intrinsic neurons in the MFC are responsible for SS in the region.     

Increased GABA levels in medial prefrontal cortex of young adults with narcolepsy

Study Objectives:

To explore absolute concentrations of brain metabolites including gamma amino-butyric acid (GABA) in the medial prefrontal cortex and basal ganglia of young adults with narcolepsy.

Design:

Proton magnetic resonance (MR) spectroscopy centered on the medial prefrontal cortex and the basal ganglia was acquired. The absolute concentrations of brain metabolites including GABA and glutamate were assessed and compared between narcoleptic patients and healthy comparison subjects.

Setting:

Sleep and Chronobiology Center at Seoul National University Hospital; A high strength 3.0 Tesla MR scanner in the Department of Radiology at Seoul National University Hospital.

Patients or Participants:

Seventeen young adults with a sole diagnosis of HLA DQB₁ 0602 positive narcolepsy with cataplexy (25.1 ± 4.6 years old) and 17 healthy comparison subjects (26.8 ± 4.8 years old).

Interventions:

N/A.

Measurements and Results:

Relative to comparison subjects, narcoleptic patients had higher GABA concentration in the medial prefrontal cortex (t = 4.10, P <0.001). Narcoleptic patients with nocturnal sleep disturbance had higher GABA concentration in the medial prefrontal cortex than those without nocturnal sleep disturbance (t = 2.45, P= 0.03), but had lower GABA concentration than comparison subjects (t = 2.30, P = 0.03).

Conclusions:

The current study reports that young adults with narcolepsy had a higher GABA concentration in the medial prefrontal cortex, which was more prominent in patients without nocturnal sleep disturbance. Our findings suggest that the medial prefrontal GABA level may be increased in narcolepsy, and the increased medial prefrontal GABA might be a compensatory mechanism to reduce nocturnal sleep disturbances in narcolepsy.

Citation:

Kim SJ; Lyoo IK; Lee YS; Sung YH; Kim HJ; Kim JH; Kim KH; Jeong DU. Increased GABA levels in medial prefrontal cortex of young adults with narcolepsy. SLEEP 2008;31(3):342-347.     

Whole-brain mapping of monosynaptic afferent inputs to the CRH neurons in the medial prefrontal cortex of mice

Corticotropin-releasing hormone (CRH) neurons are densely distributed in the medial prefrontal cortex (mPFC), which plays a crucial role in integrating and processing emotional and cognitive inputs from other brain regions. Therefore, it is important to know the neural afferent patterns of mPFC^CRH neurons, which are still unclear. Here, we utilized a rabies virus-based monosynaptic retrograde tracing system to map the presynaptic afferents of the mPFC^CRH neurons throughout the entire brain. The results show that the mPFC^CRH neurons receive inputs from three main groups of brain regions: (1) the cortex, primarily the orbital cortex, somatomotor areas, and anterior cingulate cortex; (2) the thalamus, primarily the anteromedial nucleus, mediodorsal thalamic nucleus, and central medial thalamic nucleus; and (3) other brain regions, primarily the basolateral amygdala, hippocampus, and dorsal raphe nucleus. Taken together, our results are valuable for further investigations into the roles of the mPFC^CRH neurons in normal and neurological disease states. These investigations can shed light on various aspects such as cognitive processing, emotional modulation, motivation, sociability, and pain.     

Development of brain stimulation reward in the medial prefrontal cortex: Facilitation by prior electrical stimulation of the sulcal prefrontal cortex

Electrical stimulation of the medial prefrontal cortex (MC) in rats delivered daily for seven days causes a marked improvement in the rate of acquisition of a self-stimulation response. In the present experiment, we looked at whether we could get the same facilitatory effect on self-stimulation of the MC by delivering pre-training stimulation to other points in the brain anatomically related to the MC. Electrical stimulation of the lateral hypothalamus was without effect. However, electrical stimulation of the sulcal prefrontal cortex (SC) either contralateral or ipsilateral to the MC electrode did facilitate acquisition of self-stimulation of the MC. Thus the SC and MC would appear to be part of the same substrate controlling the development of positive reinforcement in the MC.     

Involvement of medial prefrontal cortex neurons in behavioral and cardiovascular responses to contextual fear conditioning

To explore the ventral medial prefrontal cortex (vMPFC) involvement in behavioral and autonomic fear-conditioned responses to context, vMPFC synaptic transmission was temporarily inhibited by bilateral microinjections of 200 nL of the nonselective synapse blocker CoCl(2) (1 mM). Behavioral activity (freezing, motor activity and rearing) as well as evoked cardiovascular responses (arterial pressure and heart rate) was analyzed. Rats were pre-exposed to the footshock chamber (context) and shock stimulus was used unconditioned stimulus. During re-exposure to context, conditioned rats spent 80% of the session in freezing while non-conditioned rats (no shock group) spent less than 15% of the session time in freezing. Conditioned rats had significantly lower activity scores than non-conditioned animals. Exposure to context increased mean arterial pressure (MAP) and heart rate (HR) of both groups. MAP and HR of the conditioned animals were markedly increased and remained at a high and stable level, whereas MAP and HR increases in non-conditioned animals were less pronounced and declined during the session. CoCl(2) microinjected in the vMPFC significantly reduced freezing and attenuated MAP and HR increase of the conditioned group. Cobalt-induced vMPFC inhibition also significantly reduced MAP and HR increase observed in non-conditioned animals, without any behavioral changes. The effect of vMPFC acute ablation on MAP and HR did not seem to be specific to the fear response because they were also evident in non-conditioned animals. The results indicate that vMPFC integrity is crucial for expression of fear-conditioned responses to context, such as freezing and cardiovascular changes, suggesting that fear-conditioned responses to context involve cortical processing prior to amygdalar output. They also indicate a cardiovascular response observed during re-exposure of non-conditioned rats to the context is completely dependent on vMPFC integrity.     

特异性调控前额叶皮质锥体细胞活性对大鼠内脏痛行为的影响

目的:通过光遗传学手段,特异性调控内侧前额叶皮质(medial prefrontal cortex,m PFC)锥体细胞活性的变化,观察对于大鼠内脏痛及焦虑行为的影响。方法:雄性成年大鼠分为3组,随机注入AAV_(2/9)-Ca MKII-mcherry,AAV_(2/9)-Ca MKII-ChR2-mcherry以及AAV_(2/9)-Ca MKII-Arch T-mcherry病毒,特异性感染锥体细胞。病毒注射4周后,将光纤埋入双侧m PFC内,之后通过给予蓝光或黄光刺激激活含ChR2或Arch T的神经元,观察对于大鼠内脏痛和焦虑行为的影响。结果:AAV注射很好地感染了m PFC内的锥体细胞,表达ChR2的大鼠经蓝光照射后Fos表达量增高,内脏痛反应和焦虑行为受到抑制。表达Arch T的大鼠经黄光刺激后Fos表达量降低,内脏痛反应和焦虑行为增加。结论:兴奋m PFC内的锥体细胞对于内脏痛和焦虑产生明显抑制,而抑制锥体细胞则增强内脏痛和焦虑行为。     

Astrocytic cytoskeletal atrophy in the medial prefrontal cortex of a triple transgenic mouse model of Alzheimer's disease

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by the loss of cognitive functions, reflecting pathological damage to the medial prefrontal cortex (mPFC) as well as to the hippocampus and the entorhinal cortex. Astrocytes maintain the internal homeostasis of the CNS and are fundamentally involved in neuropathological processes, including AD. Here, we analysed the astrocytic cytoskeletal changes within the mPFC of a triple transgenic mouse model of AD (3 × Tg-AD) by measuring the surface area and volume of glial fibrillary acidic protein (GFAP)-positive profiles in relation to the build-up and presence of amyloid-β (Aβ), and compared the results with those found in non-transgenic control animals at different ages. 3 × Tg-AD animals showed clear astroglial cytoskeletal atrophy, which appeared at an early age (3 months; 33% and 47% decrease in GFAP-positive surface area and volume, respectively) and remained throughout the disease progression at 9, 12 and 18 months old (29% and 36%; 37% and 35%; 43% and 37%, respectively). This atrophy was independent of Aβ accumulation, as only a few GFAP-positive cells were localized around Aβ aggregates, which suggests no direct relationship with Aβ toxicity. Thus, our results indicate that the progressive reduction in astrocytic branching and domain in the mPFC can account for the integrative dysfunction leading to the cognitive deficits and memory disturbances observed in AD.     

The temporal evolution of the distribution of dendritic spines in the visual cortex of normal and dark raised mice

Summary A set of equations which define the distribution of spines along the apical dendrites have been developed. They are satisfied by the distribution of spines and its evolution with the age in the apicals of the layer V pyramidal cells of the visual cortex in normal and dark raised mice. The principal equation describes the distribution of the spines with three coefficients IF, B and K whose values have a functional relation with the age T of the animal. This relation has been defined by three additional equations whose coefficients were calculated. The equations have been used to predict the distribution of dendritic spines corresponding to age-groups of mice not previously studied and to find out the age of mice from the data of their known spine distribution resolving the inverse equations of IF (T) and B(T).     

大鼠慢性心肌梗死对前额叶皮质锥体细胞突触活性影响的电生理学研究

目的:检测大鼠满性心肌梗死所致心绞痛之后,内侧前额叶皮质(medial prefrontal cortex,mPFC)锥体细胞突触传递活性的变化,探讨调节慢性心绞痛可能的中枢机制。方法:雄性成年SD大鼠随机分为假手术组和慢性心肌梗死(chronic myocardial infarction,CMI)手术组,每组5只。手术2周后,制备mPFC区域急性脑片,通过全细胞膜片钳方法记录锥体细胞的兴奋性和抑制性突触电流,比较假手术组和手术组的突触电流活性变化。结果:相较于假手术组动物,手术组动物mPFC内兴奋性突触电流传递明显增强,抑制性突触电流传递明显减弱。同时抑制性中间神经元动作电位释放下降。结论:CMI之后mPFC内锥体细胞的兴奋性突触信号传递增强,可能是由于局部抑制性神经元活性降低所致。鉴于mPFC内锥体细胞活性对于镇痛具有正向效应,这种变化对于mPFC对慢性心绞痛的中枢抑制可能具有积极的意义。