Distribution and developmental expression of the nerve terminal protein NT75 in the rat cerebellum

Previous studies of the nerve terminal protein NT75 in the developing spinal cord have suggested an association between the appearance of NT75 immunoreactivity and the process of synaptogenesis. To examine the time course of NT75 expression further, the current study compared the localization of NT75 and the synaptic vesicle protein synaptophysin in the adult and developing rat cerebellum and in cerebellar tissue cultures. In the adult cerebellum, dense NT75 staining is confined to the molecular layer, where it is associated with parallel fiber endings of cerebellar granule cells. During development, NT75 immunoreactivity is first detectable in the cerebellar cortex as a dense band of staining in the deepest portion of the molecular layer at postnatal day 10. The stained zone expands to occupy a progressively greater portion of the molecular layer until about postnatal day 20. Synaptophysin staining occurs in granule cell processes earlier than NT75 and is found throughout the molecular layer by postnatal day 7. Quantitatively, rapid increases in both NT75 and synaptophysin occur in the first three postnatal weeks, with NT75 activity reaching levels exceeding the adult value by 50% over postnatal days 20 through 30, whereas synaptophysin plateaus at near adult levels by postnatal day 20. In cerebellar cultures, NT75 staining in neurites develops over several days, increasing coincidentally with development of synaptic contacts, whereas synaptophysin staining is already present in most neurites after only 1 day in vitro. The results indicate that NT75 expression in developing cerebellar granule cell nerve terminals is closely associated with the appearance of mature nerve terminals, suggesting that the protein may have a role in the formation/stabilization of the synaptic ending or in the mechanisms of synaptic transmission.     

Effects of denervation on neuromuscular junctions in the thyroarytenoid muscle

OBJECTIVES/HYPOTHESIS: To evaluate the effects of denervation on muscle fibers and neuromuscular junctions (NMJ) of the rat thyroarytenoid (TA) muscle with a histochemical method to monitor the status of degenerative NMJ. STUDY DESIGN: Quantitative assessment to monitor the status of degenerative muscle fibers and NMJ in the TA muscle. METHODS: Wistar rats were killed at 6, 12, 18, and 24 hours and at 2, 4, and 10 weeks after left recurrent laryngeal nerve (RLN) transection. Hematoxylin-eosin stain was used to evaluate the atrophic changes of the TA muscle. The pre- and postsynaptic structures of the NMJ were detected histochemically. These changes were evaluated by comparing the results between the treated (T) and untreated (U) sides (T/U ratio) in the same section. RESULTS: The atrophic changes in the TA muscle progressed gradually, and at 10 weeks, the T/U ratios of the entire muscle area and of the muscle fiber size decreased to 53.2 +/- 10.7% and 55.5 +/- 6.8%, respectively (P < .01). The number of nerve terminals decreased significantly at 18 hours (P < .01), and they disappeared completely by 24 hours. In contrast, at 10 weeks, 70.5 +/- 12.4% (P < .01) of acetylcholine receptors (AchRs) were preserved. CONCLUSIONS: In the rat TA muscle, denervation influences the presynaptic nerve terminals more than the postsynaptic AchRs and the muscle fibers. The results could be a basis for understanding the mechanism of laryngeal denervation and reinnervation processes in animal models.     

Localization of myosin phosphatase target subunit 1 in rat brain and in primary cultures of neuronal cells

Myosin phosphatase (PP1M) is composed of the delta isoform of the PP1 catalytic subunit (PP1cdelta), the myosin phosphatase target subunit (MYPT), and a 20 kDa subunit. Western blots detected higher amounts of the MYPT1 isoform compared to MYPT2 in whole brain extracts. The localization of MYPT1 was studied in rat brain and in primary cell cultures of neurons using specific antibodies. Analysis of lysates of brain regions for MYPT1 and PP1M by Western blots using anti-MYPT1 antibodies and by phosphatase assays with myosin as substrate suggested a ubiquitous distribution. Immunohistochemistry of tissue sections revealed that MYPT1 was distributed in all areas of the brain, with staining observed in many different cell types. Depending on the method used for fixation, the MYPT1 appeared with varying intensity in nuclei, in nucleoli, and in the cytoplasm. In primary hippocampal cultures, MYPT1 was identified by confocal microscopy in the cytoplasm and in the nucleus, whereas a predominantly cytoplasmic localization was found in cochlear nucleus cells. In cultured cells, MYPT1 and PP1cdelta colocalized with synaptophysin. PP1M activity was high in synaptosomes isolated from the cerebral cortex, but was relatively low in the postsynaptic densities. The interaction of MYPT1 with synaptophysin and with known partners (Rho-kinase, PP1cdelta) in brain extracts was shown by immunoprecipitation with anti-MYPT1. Pull-down assays from synaptosomes, using GST-MYPT1, also confirmed these interactions. In conclusion, the widespread cellular and subcellular localization of MYPT1 implies that PP1M may play an important role in the dephosphorylation of key regulatory proteins in neuronal cells.     

Synapse loss is greater in presenile than senile onset Alzheimer disease: implications for the cognitive reserve hypothesis

In the past, 'Alzheimer disease' (AD) referred to pathologic AD with clinical onset of dementia in the presenium, while 'senile dementia of the Alzheimer type' (SDAT) referred to senile onset AD. Because AD appears clinically homogeneous regardless of age of onset, the two subtypes in more recent years have not been distinguished. Pathologic differences have been noted, but synapse loss has not previously been compared between the two groups. Hypothesizing that synapse loss would be greater in presenile onset than senile onset AD, we compared synapse loss, as well as Alzheimer pathology in presenile and senile onset AD, using an ELISA method to quantify synaptophysin. Synaptophysin was significantly lower in presenile than senile AD in right frontal and bilateral parietal lobes. Neuritic plaque counts were significantly higher in presenile than senile AD in bilateral frontal and parietal lobes. Semi-quantitative evaluation of neurofibrillary tangles revealed significantly more tangles in bilateral frontal and parietal lobes in presenile than senile AD. Brain weight was significantly lower in presenile than senile AD. The differences in synapse loss and Alzheimer-type pathology in presenile and senile onset AD support the hypothesis that 'cognitive reserve' protects the human brain from neurodegenerative disease.     

运动对小鼠海马结构突触素年龄变化的影响

研究了长期适量运动（跑转笼）对Ｃ５７ＢＬ／６Ｊ小鼠海马结构突触素（ｐ３８）年龄变化的影响。以同年龄对照组心重／体重比率均值的２倍标准差作为运动有效标准，用免疫组织化学方法结合图象分析确定突触素含量。结果显示，２４月龄鼠齿状回分子层中带，海马ＣＡ３，ＣＡ１区分子层突触素免疫反应产物光密度值（ＣＯＤ值）小于１３月龄鼠（Ｐ＜０．０５，Ｐ＜０．０１），后者被检各区ＣＯＤ值小于５月龄鼠（Ｐ＜０．０５，Ｐ＜０．０１）。５月龄小鼠经过８和１９个月运动后，被检各区突触素免疫反应产物密度均较对照组显著增高（Ｐ＜０．０１）。表明从青年开始的长期适量运动能够防止增龄引起的海马结构突触素的丢失。     

急性癫痫动物模型海马突触体素的表达及意义

[目的]研究马桑内酯所致急性癫痫动物模型中海马结构内突触体素表达的变化规律,探讨突触间神经递质的传递与癫痫持续状态之间的关系。[方法]成年健康雄性SD大鼠30只,随机分为模型组及对照组,采用马桑内酯（50μg/kg体重）侧脑室注射法建立急性癫痫动物模型,应用免疫组织化学方法观察癫痫持续状态下突触体素的表达。[结果]①突触体素免疫反应产物为棕黄色点状或颗粒状沉积,在海马结构内呈明显的板层样分布。②模型组海马结构各区突触体素阳性免疫反应产物的密度较对照组普遍增加,尤以CA3区苔藓纤维层和齿状回分子层内带为甚（P﹤0.05）。[结论]突触体素表达的增强可能易化了神经递质的装载和释放,与癫痫持续状态的维持和加重相关。     

Origin of Androgen-Insensitive Poorly Differentiated Tumors in the Transgenic Adenocarcinoma of Mouse Prostate Model

Following castration, the transgenic adenocarcinoma of mouse prostate (TRAMP) model demonstrates rapid development of SV40-Tag-driven poorly differentiated tumors that express neuroendocrine cell markers. The cell population dynamics within the prostates of castrated TRAMP mice were characterized by analyzing the incorporation of 5-bromodeoxyuridine (BrdUrd) and the expression of SV40-Tag, synaptophysin, and androgen receptor (AR). Fourteen days postcastration, the remaining epithelial cells and adenocarcinoma cells were nonproliferative and lacked detectable SV40-Tag or synaptophysin expression. In contrast, morphologically distinct intraglandular foci were identified which expressed SV40-Tag, synaptophysin, and Ki67, but that lacked AR expression. These proliferative SV40-Tag and synaptophysin-expressing intraglandular foci were associated with the rare BrdUrd-retaining cells. These foci expanded rapidly in the postcastration prostate environment, in contrast to the AR- and SV40-Tag-expressing adenocarcinoma cells that lost SV40-Tag expression and underwent apoptosis after castration. Intraglandular foci of synaptophysin-expressing cells were also observed in the prostates of intact TRAMP mice at a comparable frequency; however, they did not progress to rapidly expanding tumors until much later in the life of the mice. This suggests that the foci of neuroendocrine-like cells that express SV40-Tag and synaptophysin, but lack AR, arise independent of androgen-deprivation and represent the source of the poorly differentiated tumors that are the lethal phenotype in the TRAMP model.     

吗啡对小鼠海马结构突触素表达及突触结构的影响?

目的:研究吗啡对海马结构突触素(SYN)表达的影响,为吗啡慢性中毒、成瘾和戒断症状的发生与治疗提供病理学依据.方法:将20只小鼠随机分成对照组和实验组,每组10只.对照组每只小鼠每日皮下注射生理盐水0.1 ml,实验组每只小鼠每日皮下注射吗啡0.1 ml(1 mg).注射30 d后,将两组小鼠断头处死,开颅取出脑组织并制片.切片经免疫组织化学SP法处理,光镜观察与照相,图像分析系统分析,最后行统计学处理.结果:在对照组,内嗅区、下托、齿状回的分子层和多形层及海马的始层和辐射层均显黄色,有SYN阳性表达,海马的腔隙分子层显淡黄色,有SYN弱阳性表达,锥体细胞和颗粒细胞的胞膜显黄色,呈SYN阳性表达,海马结构的平均灰度值为132.84±8.67;下托和内嗅区内有阳性表达神经元,其均值为(7.80±1.03)/mm2 .在实验组,下托和齿状回多形层显黄色,有SYN阳性表达,内嗅区及海马多形层、辐射层和腔隙分子层显深黄色,有SYN强阳性表达,锥体细胞和颗粒细胞的细胞膜显深黄色,呈SYN强阳性表达,海马结构的平均灰度值为116.27±5.70;下托和内嗅区内有强阳性表达神经元,其均值为(11.90±1.45)/mm2 .实验组的SYN阳性神经元比对照组多,SYN的表达比对照组强(P＜0.01).海马结构呈毛玻璃状,细胞染色浅,其中可见较多固缩的细胞核与空泡.结论:吗啡能促进海马结构内神经元的SYN表达和凋亡,影响突触的结构和功能.     

复明片对实验性视网膜脱离时及复位后突触素表达的影响

目的研究复明片对视网膜脱离时及复位后视网膜突触素(synaptophysin,SY)表达的影响。方法将48只家兔随机分为4组,分别为正常组、模型组、西药对照组及复明片组,采用视网膜下腔注射透明质酸钠的方法建立视网膜脱离模型,10~14d后视网膜自动复位。正常组、模型组给予生理盐水,西药对照组给予西药混合溶液,复明片组给予复明片混悬液灌胃,分别于术后1周(视网膜脱离时)及3周(视网膜复位后)采用免疫组化法观察SY表达情况。结果视网膜脱离导致SY表达显著减少,术后1周时,模型组、西药对照组及复明片组SY染色OD值分别为0.07±0.01、0.08±0.01、0.13±0.02,较正常组(0.24±0.02)明显降低(P<0.01);3周时,SY表达较视网膜脱离时增加,3组染色OD值分别为0.13±0.01、0.14±0.02、0.20±0.02,但与正常组(0.25±0.03)比较,其染色OD值仍显著降低(P<0.01;P<0.01;P<0.05);在视网膜脱离时及复位后2个时相,复明片组SY染色OD值均高于相同时间点的模型组和西药对照组,差异亦有极显著性(P<0.01)。结论视网膜脱离导致视功能严重受损,而复明片可促进视网膜神经元突触密度和分布的恢复(轴突再生),促进视觉信息的传递,从而促进视功能的恢复。