脑源性神经营养因子在早期人胚神经管发育中的定位表达

目的研究脑源性神经营养因子(BDNF)在早期人胚神经管发育过程中的定位表达。方法采用免疫细胞化学ABC法染色,研究35天人胚的发育情况。结果在人胚神经管的室带中,神经元的细胞质BDNF免疫反应阳性;在中间带,神经元的突起BDNF免疫反应阳性,一部分神经元的细胞核BDNF免疫反应阳性,另外一部分神经元的细胞核BDNF免疫反应阴性;在缘带BDNF的分布与中间带相似。神经管的头侧较尾侧BDNF阳性反应较强,神经管的腹侧BDNF阳性反应较背侧强。结论BDNF在人胚神经管免疫反应阳性,表明BDNF是诱导神经管分化发育的重要信号分子,提示BDNF在人胚神经管的发育中具有十分重要的作用。     

PV阳性神经元在孤独症模型大鼠情绪与认知相关脑区内的表达

为显示小白蛋白(Parvalbumin,PV)中间神经元在孤独症脑内的形态变化,更好地理解PV中间神经元在孤独症发病过程中的作用,本研究用免疫组化法观察了孕鼠腹腔注射丙戊酸钠(VPA)和反复冷冻刺激(RCS)法建立的两种孤独症动物模型杏仁体、前额叶皮层和海马区的PV阳性神经元的表达特点。结果显示:两种孤独症模型鼠杏仁体、前额叶皮层和海马区内的PV阳性神经元的胞体形态、大小、突起的长度和密度等与正常对照组(CTL)相比,都发生了不同程度的变化。这些结果提示,PV中间神经元在孤独症的发病过程中发挥重要的作用,它们的改变可能削弱了对孤独症相关的神经环路中锥体神经元的抑制作用。     

Wnt通路信号蛋白β-catenin和GSK-3β在孤独症模型大鼠脑中表达的变化

为探讨Wnt信号通路在孤独症发病中的作用,我们检测了Wnt信号通路中的两个关键信号蛋白分子β-catenin和GSK-3β在不同年龄阶段孤独症模型大鼠前额叶皮层,海马及小脑中的表达,同时对小脑进行了GSK-3β免疫组织化学染色检测。结果显示:孤独症模型动物的上述三个关键脑区内,β-catenin的表达水平显著升高,而GSK-3β的表达水平则明显降低;小脑内GSK-3β免疫反应阳性Purkinje细胞也明显减少。这些结果表明,孤独症模型大鼠脑内的Wnt信号通路信号传导亢进,而亢进的结果可能正是导致已知的孤独症脑内神经元发育异常的原因之一。由此提示:Wnt信号通路在孤独症的发病中起重要作用。     

小清蛋白在孤独症模型大鼠上丘中表达的年龄变化(英文)

目的:研究小清蛋白(PV)在孤独症模型大鼠上丘的表达变化。方法:用免疫组织化学及Western Blot技术,检测了胚胎期丙戊酸受药大鼠孤独症模型发育过程中上丘内PV阳性神经元的数目和PV蛋白的表达。结果:孤独症模型大鼠上丘PV阳性神经元的数目在幼年期(生后20d)明显增多,而在成年时期减少;PV蛋白的表达也相应地在幼年期增多,成年期降低(生后90d,P0.05)。结论:上丘内PV的表达随发育阶段而呈现的先增高后降低的表达变化,可能与孤独症特有的眼扫视异常有关。     

大鼠视觉中枢生后发育过程中的一氧化氮合酶阳性神经元的形态及分布变化

本文用NADPH-黄递酶组织化学技术研究了生后不同年龄段（0、7、14、21、28、35、60、120d）Wistar大鼠视皮层（17区）和上丘表层中一氧化氮合酶阳性神经元的形态及分布的变化。结果表明：视皮层17区中的一氧化氮合酶阳性神经元在生后7d时开始出现，但胞体小，树突分枝少而短，以双极细胞为主，占68.0％;14d时数量达到高峰;且14～21d中，该神经元胞体截面积明显增大，树突分枝复杂化，长度增加，多极细胞占64％;28d后一氧化氮合酶阳性神经元胞体面积，树突分枝长度明显减小，分枝简单化;35d后接近成年动物水平（120d）.生后7d时，一氧化氛合酶阳性神经元主要位于视区的白质和皮层的5、6层中，7d后该神经元及神经纤维在第2／3及4层中明显增多，这种分布状态在35d后趋于稳定。上丘表层中的一氧化氮合酶阳性细胞在生后第2周逐渐出现，第3周迅速增加，第4周达最大值，以后下降并趋于稳定水平。结果提示：大鼠生后发育过程中视觉中枢的一氧化氮合酶阳性神经元数量及形态变化与视觉发育的可塑性有关。     

以脑源性神经营养因子等干预视神经损伤大鼠视神经拉伸力学的特性分析

以拉伸力学性能指标研判以人脐血干细胞、脑源性神经营养因子干预视神经损伤动物模型的效果。以钳夹法制备视神经损伤大鼠模型,于视神经损伤大鼠模型造模7 d后,分别以人脐血干细胞、脑源性神经营养因子进行干预治疗,对各组大鼠于造模30 d后取出眶内段视神经进行组织形态观察和拉伸实验。视神经组织形态观察结果表明,视神经损伤动物模型以BDNF干预组一部分神经纤维排列较整齐,胶质细胞可见少量细胞空泡,一部分细胞核排列不规则,但视神经轴突直径改变不明显。视神经损伤动物模型以h UCBSC干预组视神经纤维排列较密集,胶质细胞核增多,多数轴突形态正常。各组动物视神经拉伸实验结果表明,视神经损伤动物模型组视神经的最大载荷、最大应力、最大应变、弹性限度载荷、弹性限度应力小于视神经损伤模型以h UCBSC、以BDNF干预组,差异显著(P0.05),视神经损伤模型以BDNF干预组的最大载荷、最大应力、最大应变、弹性限度载荷、弹性限度应力小于以h UCBSC干预组,差异显著(P0.05)。视神经损伤动物模型以BDNF和以h UCBSC干预后动物视神经的拉伸力学性能指标得到了显著提高。提示,视神经损伤模型大鼠通过BDNF、h UCBSC干预治疗后,具有明显的疗效。     

马尾神经受压大鼠脊髓圆锥神经元的形态、凋亡及脑源性神经营养因子 mRNA的表达变化

目的 观察大鼠马尾神经受压后脊髓圆锥神经元的形态变化,检测脊髓神经元凋亡数量及脑源性神经营养因子(BDNF)mRNA的表达,以及探讨上述变化的可能机制. 方法 将90只成年SD大鼠分为马尾受压模型组、假手术组与正常对照组,分别于造模术后30min、2h、4h、8h、1d、3d、1周、2周、3周取样.采用光镜与透射电镜观察马尾受压后脊髓圆锥神经细胞的形态变化;采用原位缺口末端标记(TUNEL)技术检测细胞凋亡;原位杂交法测定BDNF mRNA阳性细胞的数量.计算单位面积内的阳性细胞数,单因素方差分析比较马尾受压后不同时间组与对照组之间的差异. 结果 马尾神经受压可导致脊髓圆锥神经元形态结构出现明显伤害性改变.TUNEL染色与BDNF mRNA原位杂交显示,阳性细胞分别于制模术后8h、4h起较对照组有明显增多,并于术后3d达高峰,术后3周上述阳性细胞的数量仍明显高于对照组. 结论 马尾神经受压后,可导致相应脊髓圆锥神经细胞结构的明显变化,其凋亡数量明显增加,说明马尾受损可导致中枢神经元的不可逆损伤,这可能是马尾综合征逾期手术效果不理想的重要原因之一.在马尾严重受压后,神经元及胶质细胞的BDNFmRNA的表达明显增加,可能对神经细胞的保护及修复起一定作用.     

慢性脑缺血大鼠海马CA_1区中的BDNF表达

为了观察慢性脑缺血时海马CA1区中BDNF的表达变化,探讨缺血性脑损伤及修复机制。我们将Wistar大鼠双侧颈总动脉结扎制成慢性脑缺血动物模型。分为三组:正常对照组、慢性脑缺血30d和120d组。分别于术后30d和120d处死动物,行BDNF免疫组化染色,计数各组大鼠海马CA1区中的阳性神经元数。结果显示:(1)在对照组的海马CA1区可见BDNF较强的表达;(2)在慢性脑缺血30d时,海马CA1区BDNF的表达高于缺血120d组及对照组(P<0.05)。而120d时,海马CA1区BDNF的表达与对照组无显著性差异(P>0.05)。结果提示:(1)在正常大鼠海马CA1区有BDNF表达,能维持神经元的存活;(2)慢性脑缺血时,BDNF在海马CA1区的表达先升后降。     

大鼠海马CA1和CA3区发育过程中酪氨酸激酶受体B的分布

为了观察脑源性神经营养因子的特异性受体—酪氨酸激酶受体 B在发育过程中的大鼠海马 CA1和 CA3区的分布 ,本研究使用免疫组织化学 A BC法研究了生后几个时间段的酪氨酸激酶受体 B的分布特点。结果表明 :其免疫阳性产物仅出现在神经元胞体中且只位于胞浆 ,胶质细胞中未见分布。生后零天组偶见免疫阳性细胞 ;生后 5天组免疫阳性细胞较生后 0天组明显增多 ,但分布也尚较少 ;生后 10、15、2 0、30天 4组免疫阳性细胞呈明显的逐渐增多趋势。成年组的免疫阳性细胞与生后 30天组无明显差别。本实验结果提示 :发育早期大鼠海马 CA1和 CA 3区锥体细胞产生酪氨酸激酶受体 B,并通过对它的分泌调节 ,控制脑源性神经营养因子对神经元的作用 (突触的发生、发育、维持及神经元损伤后修复等     

BDNF及其受体TrkB在成年恒河猴脊髓的表达

目的观察脑源性神经营养因子BDNF及其高亲合力受体TrkB在成年恒河猴胸髓的表达分布,为成年恒河猴脊髓BDNF、TrkB的功能研究提供形态学依据。方法灌注固定成年雄性恒河猴,取其胸段脊髓制作冰冻切片,用BDNF、TrkB特异性抗体行免疫组织化学SP法染色,观察BDNF、TrkB免疫阳性物质在胸段脊髓的分布。结果成年恒河猴胸髓内均可见BDNF、TrkB免疫反应阳性神经元,反应产物呈棕黄色颗粒。BDNF和TrkB免疫反应阳性细胞遍及胸髓灰质各板层,在腹角、侧角以及中央管邻近区域均可见阳性神经元,它们的阳性产物主要定位于神经元胞浆中,胞浆着色深,胞浆与胞核的界线清楚。BDNF免疫反应细胞的胞核未见染色,而不同的TrkB阳性细胞其胞核染色不一。结论成年恒河猴胸髓存在BDNF、TrkB的表达,其作用可能与猴脊髓的正常生理功能和损伤后的修复有关。     

Expression of brain-derived neurotrophic factor in rat dorsal root ganglia, spinal cord and gracile nuclei in experimental models of neuropathic pain.

Chronic constriction injury of the sciatic nerve and lumbar L5 and L6 spinal nerve ligation provide animal models for pain syndromes accompanying peripheral nerve injury and disease. In the present study, we evaluated changes in brain-derived neurotrophic factor (BDNF) immunoreactivity in the rat L4 and L5 dorsal root ganglia (DRG) and areas where afferents from the DRG terminates (the L4/5 spinal cord and gracile nuclei) in these experimental models of neuropathic pain. Chronic constriction injury induced significant increase in the percentage of small, medium and large BDNF-immunoreactive neurons in the ipsilateral L4 and L5 DRG. Following spinal nerve ligation, the percentage of large BDNF-immunoreactive neurons increased significantly, and that of small BDNF-immunoreactive neurons decreased markedly in the ipsilateral L5 DRG, while that of BDNF-immunoreactive L4 DRG neurons of all sizes showed marked increase. Both chronic constriction injury and spinal nerve ligation induced significant increase in the number of BDNF-immunoreactive axonal fibers in the superficial and deeper laminae of the L4/5 dorsal horn and the gracile nuclei on the ipsilateral side.Considering that BDNF may modulate nociceptive sensory inputs and that injection of antiserum to BDNF significantly reduces the sympathetic sprouting in the DRG and allodynic response following sciatic nerve injury, our results also may suggest that endogenous BDNF plays an important role in the induction of neuropathic pain after chronic constriction injury and spinal nerve ligation. In addition, the increase of BDNF in L4 DRG may contribute to evoked pain which is known to be mediated by input from intact afferent from L4 DRG following L5 and L6 spinal nerve ligation.     

Olfactory sensory deprivation increases the number of proBDNF-immunoreactive mitral cells in the olfactory bulb of mice

In the olfactory bulb, apoptotic cell-death induced by sensory deprivation is restricted to interneurons in the glomerular and granule cell layers, and to a lesser extent in the external plexiform layer, whereas mitral cells do not typically undergo apoptosis. With the goal to understand whether brain-derived neurotrophic factor (BDNF) mediates mitral cell survival, we performed unilateral naris occlusion on mice at postnatal day one (P1) and examined the subsequent BDNF-immunoreactive (BDNF-ir) profile of the olfactory bulb at P20, P30, and P40. Ipsilateral to the naris occlusion, there was a significant increase in the number of BDNF-ir mitral cells per unit area that was independent of the duration of the sensory deprivation induced by occlusion. The number of BDNF-ir juxtaglomerular cells per unit area, however, was clearly diminished. Western blot analysis revealed the presence of primarily proBDNF in the olfactory bulb. These data provide evidence for a neurotrophic role of proBDNF in the olfactory system of mice and suggest that proBDNF may act to protect mitral cells from the effects of apoptotic changes induced by odor sensory deprivation.     

The morphology and distribution of peptide-containing neurons in the adult and developing visual cortex of the rat. IV. Avian pancreatic polypeptide

Summary Immunocytochemical techniques were used to investigate the morphology and distribution of avian pancreatic polypeptide-like immunoreactive neurons in the visual cortex of albino rats at various ages from the first postnatal day to adulthood. In the adult, immunoreactive neurons were located in layers II to VI but were somewhat concentrated in the deeper cortical layers. The overwhelming majority of labelled cells exhibited morphologies characteristic of multipolar, bitufted and bipolar varieties of non-pyramidal neurons as described in Golgi preparations of rat visual cortex. However, a few immunoreactive pyramidal neurons were also observed.On the first postnatal day, a small number of immature non-pyramidal neurons were observed in the subplate region. Labelled cells appeared in the more superficial layers at the beginning of the second postnatal week and attained a distribution similar to that observed in adult animals during the third week. The morphological maturation of immunoreactive neurons occurred gradually during the first two postnatal weeks and at day 21, they appeared qualitatively indistinguishable from their adult counterparts.     

Cytidine-5-diphosphocholine supplement in early life induces stable increase in dendritic complexity of neurons in the somatosensory cortex of adult rats

Cytidine-5-diphosphocholine (CDP-choline or citicholine) is an essential molecule that is required for biosynthesis of cell membranes. In adult humans it is used as a memory-enhancing drug for treatment of age-related dementia and cerebrovascular conditions. However the effect of CDP-choline on perinatal brain is not known. We administered CDP-choline to Long Evans rats each day from conception (maternal ingestion) to postnatal day 60 (P60). Pyramidal neurons from supragranular layers 2/3, granular layer 4 and infragranular layer 5 of somatosensory cortex were examined with Golgi-Cox staining at P240. CDP-choline treatment significantly increased length and branch points of apical and basal dendrites. Sholl analysis shows that the complexity of apical and basal dendrites of neurons is maximal in layers 2/3 and layer 5. In layer 4 significant increases were seen in basilar dendritic arborization. CDP-choline did not increase the number of primary basal dendrites on neurons in the somatosensory cortex. Primary cultures from somatosensory cortex were treated with CDP-choline to test its effect on neuronal survival. CDP-choline treatment neither enhanced the survival of neurons in culture nor increased the number of neurites. However significant increases in neurite length, branch points and total area occupied by the neurons were observed. We conclude that exogenous supplementation of CDP-choline during development causes stable changes in neuronal morphology. Significant increase in dendritic growth and branching of pyramidal neurons from the somatosensory cortex resulted in enlarging the surface area occupied by the neurons which we speculate will augment processing of sensory information.     

脑源性神经营养因子对出生后大鼠周围神经髓鞘化的作用

目的:探讨脑源性神经营养因子(BDNF)对出生后大鼠周围神经髓鞘化的作用及其作用机制.方法:用光镜和电镜方法检测坐骨神经纤维的髓鞘化情况;用免疫印迹方法检测坐骨神经p75的表达;用免疫荧光方法检测坐骨神经雪旺细胞内核转录因子(NF-kB)的表达.结果:出生后3 d,与对照组相比,实验组坐骨神经髓鞘化的神经纤维数目减少,p75表达下调,雪旺细胞内NF-KB转移率下调;出生后14 d,与对照组相比,实验组坐骨神经出现较多的髓鞘化异常.结论:内源性BDNF影响出生后大鼠周围神经的髓鞘化,尤其在早期作用明显;内源性BDNF主要通过p75和NF-kB信号途径调节出生后大鼠周围神经的髓鞘化.     

Apoptosis and retinal projections in the dorsal lateral geniculate nucleus after monocular deprivation during the later phase of the critical period in the rat

The visual cortex in the rat is matured physiologically by postnatal day 30, but the visual system retains the potential to be reorganized until postnatal day 45. Therefore, we defined the period from postnatal days 28-45 as the ‘late critical phase’. To examine whether monocular deprivation during the late critical phase gives rise to neuronal apoptosis in the dorsal lateral geniculate nucleus (dLGN), we used the terminal deoxyribo-nucleotidyl transferase-mediated dUTP-digoxigenin nick end-labeling method and anterograde tracing. The number of apoptotic cells in the dLGN after monocular deprivation at postnatal day 28 showed little difference from control at postnatal day 29, but was significantly increased to more than fourfold of control ipsilaterally to the monocular deprivation at postnatal day 35 and to more than 10-fold of control bilaterally at postnatal day 40. In the control, there were almost no apoptotic neurons in the dLGN on either side at postnatal day 40. In the nucleus ipsilateral to the monocular deprivation, approximately half the apoptotic neurons were found in an area that did not receive a retinal projection. These findings suggest that the biological process of increased apoptosis in the dLGN of rats that received monocular deprivation in the late critical phase may be different from that in the early critical phase. The increased number of apoptotic cells in the dLGN in the late critical phase may not be simply the result of monocular deprivation.     

Expression of brain-derived neurotrophic factor in the rat forebrain and upper brain stem during postnatal development: an immunohistochemical study

The present study was undertaken to characterize the regional and temporal patterns of brain-derived neurotrophic factor (BDNF) in the rat forebrain and upper brain stem during postnatal development using an immunohistochemical approach. Results indicated that BDNF-immunoreactive (IR) cells could be divided into three groups based on their postnatal developmental patterns: (group 1) BDNF-IR cells were first detected between postnatal days (PND) 1 and 7, and thereafter they increased in number and remained stable during later stages of ontogeny; (group 2) BDNF-IR cells progressively increased in number with age, and then decreased in adults; (group 3) numerous BDNF-IR cells detected between PND 1 and 7 showed a dramatic reductions in number with few IR cells in adults. In contrast, the developmental pattern of most BDNF-IR fibers differed from that of IR neurons, i.e. they appeared between PND 1-28 and thereafter continued to increase in number showing a maximum level in adults. Additionally, BDNF-IR cells in the superficial layer of the neocortex and IR fibers in the stratum oriens of CA2 first appeared as late as PND 28 and in adults, respectively. After colchicine treatment, reexpression or a marked increase in the number of BDNF-IR neurons was observed in many areas of the adult brain where a progressive decrease in BDNF-IR cell numbers during development and scant or some IR neurons in adults were shown. These results showed both transient and persistent expression of BDNF in various regions of the developing rat brain.     

Effects of hyperthermic seizures on the developing primary visual cortex of the rat

Febrile seizures occurring during childhood have been shown to interfere with the development of cognitive functions. However, an alteration of the developing sensory systems might also result from febrile seizures. In order to test this hypothesis, seizures were induced by hyperthermia in Long Evans rats on postnatal day 10. Extracellular single neuron recordings were carried out from postnatal days 15 to 30 and at adulthood. The response of neurons in the primary visual cortex to drifting sinusoidal gratings was recorded in anaesthetized rats. As soon as postnatal day 15, the neurons of rats having experienced a hyperthermic seizure showed significantly lower optimal spatial frequencies (SF), broader directional and temporal bandwidths, as well as higher contrast thresholds than did neurons recorded in normal rats. At adulthood, significantly broader spatial bandwidths and lower optimal temporal frequencies (TF) were obtained from neurons of rats subjected to hyperthermia. These results suggest that febrile seizures during infancy could affect the development of spatio-temporal receptive field properties of neurons in primary visual cortex. Such alterations of a sensory system might contribute to the cognitive deficits associated with early-onset febrile seizures.     

节制饮食增强猫视皮层γ-氨基丁酸和脑源性神经营养因子的表达

目的探讨节制饮食对猫初级视皮层内抑制性神经递质γ-氨基丁酸(GABA)和脑源性神经营养因子(BDNF)表达的影响。方法 Nissl染色示初级视皮层分层并用于神经元计数;免疫组织化学方法标记6只猫GABA和BDNF免疫阳性神经元。切片于Olympus光学显微镜下观察摄片,用Image-Pro Express 6.0软件进行细胞计数和免疫阳性反应的吸光度值统计。结果与正常饮食对照猫相比,节制饮食组猫的初级视皮层神经元平均密度无显著变化,GABA免疫阳性神经元密度显著增大,阳性反应的平均吸光度值显著升高;BDNF免疫阳性细胞的密度和阳性反应的平均吸光度值亦明显增大。结论节制饮食能显著增强猫视皮层内GABA和BDNF的表达,这可能对视皮层神经元的结构和功能有保护作用,从而延缓衰老进程。