Developmental Changes in Levels of Translatable mRNAs for Enolase Isozymes in Chicken Brain

Using chicken brain mRNAs, alpha and gamma enolase precursors were synthesized in the rabbit reticulocyte cell-free translation system. The product proteins showed molecular weights almost identical to those of the mature subunits. The levels of translatable mRNAs for alpha and gamma subunits were determined by the cell-free translation system and immunoprecipitation with specific antisera, during development of chicken brain. The level of alpha mRNA was high at any developmental stage of the brain. On the other hand, the gamma mRNA level was very low at the early embryonic stage, and increased rapidly during development of the brain. These changes were closely correlated with those of the corresponding enzyme activities, indicating that the levels of enolase activities in developing brain were controlled primarily by the level of the translatable alpha and gamma mRNAs.     

神经生长因子在不同周龄小鼠睾丸组织中的表达

目的研究神经生长因子在小鼠不同周龄睾丸组织中的定量和定位表达。方法分别剖取不同周龄雄性小鼠的睾丸组织,部分提取总RNA,real-time PCR相对定量分析神经生长因子mRNA的表达量;另外部分组织固定、包埋,进行SABC法免疫组化分析,以观察神经生长因子蛋白在各周睾丸组织中的定位。结果Real-timePCR定量分析表明:小鼠生后1周龄睾丸组织有神经生长因子mRNA的表达,生后3周龄表达量达峰值,5周之后随鼠龄的增加呈下降趋势,成年小鼠睾丸组织的神经生长因子mRNA表达维持在一定水平。免疫组化定位分析显示:睾丸组织的神经生长因子蛋白表达于小鼠出生后的各个时期内,1周龄睾丸组织免疫阳性反应主要位于支持细胞,精原细胞也有着色;3周龄睾丸组织的间质细胞、各级生精细胞、支持细胞、管周肌样细胞表达均呈现阳性;5周后的睾丸组织内神经生长因子呈低水平表达,主要表达于间质细胞和生精细胞内。结论神经生长因子mRNA的表达量随着小鼠睾丸的生长发育期存在着一定的规律性变化;神经生长因子蛋白的表达在小鼠睾丸生长发育的不同时期其主要表达部位不同。     

Age-Related Changes in the DNA and RNA Content of the Brain in Spontaneously Hypertensive Rats

To elucidate the effects of aging accompanied with hypertension on brain nucleic acid, we measured both the DNA and RNA contents of six specific brain regions in adult (5–6 months old) and aged (18–22 months old) female spontaneously hypertensive rats (SHRs). Although no statistical difference was observed in the RNA content, the DNA content did tend to increase in the hippocampal CA1 of aged SHR (4.24 ± 0.55 ng/g protein, mean ± SD, n = 6) in comparison to that of adult SHR (3.21 ± 0.71 ng/g protein, n = 4). Hence, aged SHRs showed a significant decrease in the RNA to DNA ratio in the CA1 subfield of the hippocampus (3.79 ± 0.61) compared to adult SHR (5.27 ± 0.81). On the other hand, no other regions, except for the dorsolateral region of the striatum, showed any difference in the RNA/DNA ratio between aged and adult SHR. We therefore conclude that subtle changes in the nucleic acid occur in vulnerable regions of the brain in aged SHRs.     

bFGF、NGF、EGF及其受体在人胚神经管早期发育中的表达

研究bFGF、NGF、EGF及其受体在人胚神经管早期发育中的表达。方法 应用免疫组织化学方法和图像分析。结果显示bFGF和NGF的表达时序不同,bFGF阳性细胞出现较早,在所检测在各个发育阶段均呈阳性表达,而NGF出现较晚,随着胚龄增加,免疫阳性着色逐渐增强,bFGF分布较NGF广泛,而EGF在所检测的各个发育阶段均呈阴性。flg、TrkA、EGFR表达时序和分布相似,三者在所检测的各个发育阶段均阳性。结果表明NGF和bFGF均通过其特异性受体介导,在胚胎神经管形成和分化的不同阶段发挥着重作用,EGF及其受体的作用有待进一步研究。     

Nerve Growth Factor Administration Attenuates Cognitive but Not Neurobehavioral Motor Dysfunction or Hippocampal Cell Loss Following Fluid-Percussion Brain Injury in Rats

Abstract: Lateral fluid-percussion brain injury in rats results in cognitive deficits, motor dysfunction, and selective hippocampal cell loss. Neurotrophic factors have been shown to have potential therapeutic applications in neurodegenerative diseases, and nerve growth factor (NGF) has been shown to be neuroprotective in models of excitotoxicity. This study evaluated the neuroprotective efficacy of intracerebral NGF infusion after traumatic brain injury. Male Sprague-Dawley rats received lateral fluid-percussion brain injury of moderate severity (2.1–2.3 atm). A miniosmotic pump was implanted 24 h after injury to infuse NGF (n = 34) or vehicle (n = 16) directly into the region of maximal cortical injury. Infusions of NGF continued until the animal was killed at 72 h, 1 week, or 2 weeks after injury. Animals were evaluated for cognitive dysfunction (Morris Water Maze) and regional neuronal cell loss (Nissl staining) at each of the three time points. Animals surviving for 1 or 2 weeks were also evaluated for neurobehavioral motor function. Although an improvement in memory scores was not observed at 72 h after injury, animals receiving NGF infusions showed significantly improved memory scores when tested at 1 or 2 weeks after injury compared with injured animals receiving vehicle infusions ( p < 0.05). Motor scores and CA3 hippocampal cell loss were not significantly different in any group of NGF-treated animals when compared with controls. These data suggest that NGF administration, in the acute, posttraumatic period following fluid-percussion brain injury, may have potential in improving post-traumatic cognitive deficits.     

原花青素对脑缺血再灌损伤大鼠模型的影响

目的研究原花青素对脑缺血/再灌损伤(ischemia/reperfusion,I/R)大鼠神经功能评分(neurologicaldeficit score,NDS)、脑梗死体积、脑含水量等指标的药理作用。方法采用大鼠大脑中动脉阻断(middle cerebralartery occlusion,MCAO)法复制类似人类缺血性卒中的I/R损伤模型。结果该模型各时间点内均有程度不同的神经功能缺失,原花青素给药组神经功能评分明显低于对照组(P0.05),假手术组大鼠均无神经功能缺失,脑水肿情况均较对照组明显改善(P0.05),脑梗死体积与盐水对照组相比差异有显著性(P0.05),而假手术组均未见有梗死灶。结论原花青素具有一定的保护大鼠I/R后受损脑组织的作用,可供后续研究,并可为缺血性卒中使用原花青素治疗提供确凿的理论依据。     

神经生长因子生物活性检定方法的建立

采用鸡胚背根神经节体外培养法,建立了神经生长因子生物活性的检定方法,对判定神经生长因子的生物活性单位制定了明确统一的判定标准     

Catecholamine Content Changes in Brain Regions of Spontaneously Hypertensive Rats Under Immobilization Stress

We compared the effect of immobilization stress on noradrenaline (NA) and 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG) content in two brain regions--diencephalon and pons-medulla oblongata--in young and adult spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). In SHR, NA content decreased with time after the onset of the stress, whereas levels of its metabolite MHPG increased. In WKY, NA and MHPG showed no change. The MHPG/NA ratio in both regions increased relative to the duration of the stress in SHR, whereas it remained almost constant in WKY. The rate of increase in the ratio was much higher in the diencephalon of adult (12-week-old) than of young (4-week-old) SHR. In SHR, NA turnover in the brain is readily affected by environmental stress, and these changes in the noradrenergic system may induce or sustain hypertension.     

花生油对2型糖尿病大鼠海马CA1区神经生长因子及胆碱乙酰转移酶表达的影响

目的通过观察2型糖尿病大鼠海马CA1区神经生长因子（NGF）和胆碱乙酰转移酶（ChAT）表达的改变,研究花生油对2型糖尿病大鼠海马神经元NGF及ChAT表达的影响,探讨花生油在防治糖尿病脑病中的作用。方法 60只健康雄性SD大鼠随机分为4组：正常对照组（C组）、2型糖尿病组（T2DM组）、2型糖尿病给予2 mL花生油组（T2DM＋2 mL组）及2型糖尿病给予5 mL花生油组（T2DM＋5 mL组）。其中C组给予正常饮食,糖尿病组大鼠给予高脂饮食喂养,2个月后,按25 mg/kg体质量腹腔注射链脲佐菌素（STZ）制成2型糖尿病模型,T2DM组、T2DM＋2 mL组及T2DM＋5 mL组大鼠继续给予高脂饮食。糖尿病造模1个月后处死全部大鼠,行脑冰冻切片,用免疫组织化学方法检测各组大鼠海马CA1区NGF和ChAT的表达。结果 （1）T2DM组大鼠海马CA1区NGF表达比C组明显降低（P〈0.05）,T2DM＋2 mL组及T2DM＋5 mL组大鼠海马CA1区NGF表达均明显高于未给予花生油的T2DM组（P〈0.05）。（2）T2DM组大鼠海马CA1区ChAT表达显著低于C组（P〈0.05）,T2DM＋2 mL组和T2DM＋5 mL组大鼠海马CA1区ChAT表达均明显高于未给予花生油的T2DM组（P〈0.05）。结论 2型糖尿病大鼠海马CA1区神经生长因子表达降低,胆碱能神经元数量减少,这可能是2型糖尿病脑病发生的原因之一。花生油能增加2型糖尿病大鼠海马区内神经生长因子表达,促进胆碱能神经元存活,表明花生油具有一定的保护大鼠糖尿病脑病的作用。     

Developmental Changes in Enzymes Involved in Dolichyl Phosphate Metabolism in Cultured Embryonic Rat Brain Cells

The rates of synthesis of dolichol-linked oligosaccharide intermediates and protein N-glycosylation increased substantially during a developmental period corresponding to glial differentiation in primary cultures of embryonic rat brain. In this study developmental changes in three enzymes involved in dolichyl phosphate (Dol-P) metabolism have been examined by in vitro assays and correlated with the induction pattern for lipid intermediate synthesis and protein N-glycosylation. Dolichyl pyrophosphate (Dol-P-P) phosphatase activity was relatively low during the first 9 days in culture, but it increased significantly between days 9 and 25. Dol-P-P phosphatase did not change appreciably between days 22 and 30 in culture. A kinetic analysis of the developmental change in Dol-P-P phosphatase activity revealed that the Vmax increased 10-fold between days 4 and 22, and there was also a significant change in the apparent Km for Dol-P-P. Dolichol kinase activity increased during the period (9-15 days) when there was a significant induction in oligosaccharide-lipid synthesis and protein N-glycosylation, and then declined in parallel with lipid intermediate synthesis and protein N-glycosylation. Dol-P phosphatase activity was present at relatively low levels for the first 9 days in culture, but it increased steadily between days 9 and 30. A kinetic comparison of the activity in membrane fractions from brain cells cultured for 9 and 25 days indicated that there was a 10-fold increase in enzyme protein with unaltered affinity for Dol-P.(ABSTRACT TRUNCATED AT 250 WORDS)     

Protein Kinase C in Rat Brain Is Altered by Developmental Lead Exposure

The absence of learning-related redistribution of hippocampal protein kinase C (PKC) has been correlated with impairment of learning performance induced by developmental lead (Pb) exposure. This study was designed to examine whether the properties of brain PKC are altered by chronic Pb exposure during development. Two-tenth percent Pb acetate was administered to pregnant and lactating dams and then administered to weanlings in drinking water until postnatal day (PN) 56. Effects of Pb on translocation of PKC were studied in brain slices prepared from hippocampus. When the slices were treated with 0.33 M phorbol-12, 13-dibutyrate (PDBu) for 15 min, a significant increase in PKC activity was observed in the membrane fraction of hippocampal slices from Pb-exposed rats, suggesting that chronic Pb exposure potentiates PDBu-activated PKC translocation. Data obtained from saturation binding assays in the frontal cortices of Pb-exposed rats showed a decrease in the dissociation constant (K_D) in both membrane and cytosolic PKC. A decrease in the total binding sites (B_max) of [³H]PDBu binding was only observed in membrane PKC. Furthermore, developmental Pb exposure decreased PKC-, but not PKC-, -II, and - in the membrane fraction of the hippocampus and the frontal cortex. These results indicate that chronic Pb exposure during development increases phorbol ester binding affinity, enhances phorbol ester-induced translocation of PKC, and down-regulates membrane PKC, mainly PKC-.     

Triptolide Upregulates NGF Synthesis in Rat Astrocyte Cultures

Triptolide (T10), an extract from the traditional Chinese herb, Tripterygium wilfordii Hook F (TWHF), has been shown to attenuate the rotational behavior induced by d-amphetamine and prevent the loss of dopaminergic neurons in the substantia nigra in rat models of Parkinson’s disease. To examine if the neuroprotective effect is mediated by its stimulation of production of neurotrophic factors from astrocytes, we investigated the effect of T10 on synthesis and release of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) in rat astrocyte cultures. T10 did not affect the synthesis and release of either BDNF or GDNF. However, it significantly increased NGF mRNA expression. It also increased both intracellular NGF and NGF level in culture medium. These results indicate that the neuroprotective effect of T10 might be mediated, at least in part, via a stimulation of the production and release of NGF in astrocytes. Authors Bing Xue and Jian Jiao contributed equally to this work.     

神经营养因子NGF、BDNF与围绝经期妇女认知功能关系的研究进展

神经营养因子(NTFs)是近几年神经科学研究的热点,研究显示它在神经系统中发挥独特的作用,尤其是神经生长因子(NGF)、脑源性神经营养因子(BDNF)在脑内功能及其表达调控方面具有重要作用。围绝经期妇女随着雌激素水平的降低会产生认知功能的减退,有研究发现去卵巢动物(OVX)雌激素水平降低可以导致某些NGF、BDNF的丢失。通过启动内源性NGF和BDNF的表达而实现对神经元的保护可能为雌激素替代治疗(ERT)脑保护作用的一种机制。本文就近几年的研究进展做一简要综述。     

蛇毒神经生长因子诱导PC12细胞分化超微结构的观察

目的观察蛇毒神经生长因子（Nerve growth factor,NGF）诱导大鼠肾上腺嗜铬细胞瘤细胞系（Pheo—chromocytoma cells。PC12）细胞分化后,细胞超微结构的改变。方法取对数生长期PC12细胞接种24孔板,设200ng／ml NGF实验组和对照组。培养72h,离心,分别收集细胞制成电镜标本,镜下观察各组细胞超微结构的改变。结果与对照组细胞相比,实验组细胞长出大量突起．并且胞质的细胞器逐渐消失．出现较多的脂滴。结论广西眼镜蛇毒神经生长因子可以促进PC12细胞增殖．并诱导其向神经样细胞分化,长出突触。     

Immunocytochemical localization of nerve growth factor (NGF) in the submandibular gland of adult mice by light and electron microscopy

Summary Nerve growth factor (NGF) was localized in the submandibular gland of adult male mice by a direct immunocytochemical method using highly purified antibodies against NGF coupled to horseradish peroxidase. In light microscopic sections the reaction product was entirely confined to the cells of the secretory tubules. The acinar part of the gland was free of reaction product. This finding was confirmed by electron microscopy. Within the cells NGF was localized exclusively in the apical secretory granules.No reaction was observed in the rough endoplasmic reticulum, the Golgi region or in the granules of the basal part of the cells. This observation favours the assumption that NGF is derived from a precursor molecule and that the precursor is transformed into immunologically active NGF within the secretory granules during their transport from the basal to the apical part of the tubular cells. Stimulation of the submandibular gland with carbachol (2 mg/kg) led to a massive release of the content of the secretory granules, including NGF, into the salivary duct.We wish to thank Dr. C. Rufener, Geneva, for communicating to us a new method of antibodyperoxidase coupling and Mrs. M. Durand-Wenger for her excellent technical assistance. This study was supported by the Swiss National Foundation for Scientific Research (Grant Nr. 3.432.74)     

Brain Monoamine Changes in Rats After Short Periods of Ozone Exposure

We have shown in our laboratory that cat's and rat's sleep disturbances are produced by 24 h of ozone (O₃) exposure, indicating that the central nervous system is affected by this gas. To demonstrate the probable changes in brain neurotransmitters, we evaluated the monoamine contents of the midbrain and striatum of rats exposed to 1 part per million O₃ for 1 or 3 hours periods. The results were compared with rats exposed to fresh air and to those exposed to 3 hours of O₃ followed by 1 or 3 hours of fresh air. We found a significant increase in dopamine (DA) and its metabolites noradrenaline (NA) and 3,4 dihydroxyphenylacetic acid (DOPAC), as well as an increase in the 5-hydroxyindolacetic acid (5-HIAA) contents of the striatum. There were no changes in homovanillic acid (HVA) and serotonin (5-HT) levels during O₃ exposure. Additionally, an increase in DA, NA and 5-HIAA in the midbrain during O₃ exposure was observed. Turnover analysis revealed that DA increased more than its metabolites in both the midbrain and striatum. However, the metabolite of 5-HT, i.e. 5-HIAA, increased more than its precursor, this reaching statistical significance only in the midbrain. These findings demonstrate that O₃ or its reaction products affect the metabolism of major neurotransmitter systems as rapidly as after 1 h of exposition.     

Vanadyl Sulfate Administration Protects the Streptozotocin-Induced Oxidative Damage to Brain Tissue in Rats

Diabetes mellitus manifests itself in a wide variety of complications and the symptoms of the disease are multifactorial. The present study was carried out to investigate the effects of vanadyl sulfate on biochemical parameters, enzyme activities and brain lipid peroxidation, glutathione and nonenzymatic glycosylation of normal- and streptozotocin-diabetic rats. Streptozotocin (STZ) was administered as a single dose (65 mg/kg) to induce diabetes. A dose of 100 mg/kg vanadyl sulfate was orally administered daily to STZ-diabetic and normal rats, separately until the end of the experiment, at day 60. In STZ-diabetic group, blood glucose, serum sialic and uric acid levels, serum catalase (CAT) and lactate dehydrogenase (LDH) activities, brain lipid peroxidation (LPO) and nonenzymatic glycosylation (NEG) increased, while brain glutathione (GSH) level and body weight decreased. In the diabetic group given vanadyl sulfate, blood glucose, serum sialic and uric acid levels, serum CAT and LDH activities and brain LPO and NEG levels decreased, but brain GSH and body weight increased.The present study showed that vanadyl sulfate exerted antioxidant effects and consequently may prevent brain damage caused by streptozotocin-induced diabetes.     

Treadmill exercise after social isolation increases the levels of NGF,BDNF, and synapsin I to induce survival of neurons in the hippocampus,and improves depression-like behavior

[Purpose]

We investigated the effects of 8 weeks of treadmill exercise on nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and synapsin I protein expression and on the number of 5-bromo-2''-deoxyuridine-5''-mono-phosphate (BrdU)-positive cells in the dentate gyrus of the hippocampus in socially isolated rats. Additionally, we examined the effects of exercise on the number of serotonin (5-HT)- and tryptophan hydroxylase (TPH)-positive cells in the raphe nuclei and on depression behaviors induced by social isolation.

[Methods]

Forty male Sprague-Dawley rats were divided into four groups: (1) group housing and control group (GCG, n = 10); (2) group housing and exercise group (GEG, n = 10); (3) isolated housing and control group (ICG, n = 10); and (4) isolated housing and exercise group (IEG, n = 10). After 1 week of housing under the normal condition of 3 animals per cage, rats were socially isolated via transfer to individual cages for 8 weeks. Rats were then subjected to treadmill exercise for 5 days per week for 8 weeks during which time the speed of the treadmill was gradually increased.

[Results]

Compared to the GCG, levels of NGF, BDNF, and synapsin I were significantly decreased in the ICG and significantly increased in the IEG (p < 0.001 respectively). Significantly more BrdU-positive cells in the GEG were present as compared to the GCG and ICG, and more BrdU-positive cells were found in the IEG as compared to the ICG (p < 0.001). 5-HT-positive cells in the GEG were significantly increased compared to the GCG and ICG, and more of these cells were found in the IEG as compared to the ICG (p < 0.01). TPH-positive cells in the GEG were significantly increased compared to those in the GCG and ICG (p < 0.05). In the forced swim test, immobility time was significantly increased in the ICG and significantly decreased in the IEG as compared to the ICG (p < 0.01).

[Conclusion]

These results showed that regular treadmill exercise following social isolation not only increased the levels of NGF, BDNF, and synapsin I to induce survival of neurons in the hippocampus but also improved depression by increasing the number of serotonergic cells in the raphe nuclei.     

Involvement of Nerve Growth Factor and Its Receptor in the Regulation of the Cholinergic Function in Aged Rats

The role of nerve growth factor (NGF) and its receptor (NGFR) in the regulation of cholinergic activity has been studied during the aging process. NGFRs were quantified in cortical membranes using a radioactive binding assay. NGF levels and choline acetyltransferase (ChAT) activity were determined in cortex, hippocampus, neostriatum, and septum. These assays were performed in both adult (6-month-old) and aged (36-month-old) rats. High- and low-affinity 125I-NGF binding sites were present in cortex of adult and aged rats. Furthermore, we observed a decrease in number and affinity of both NGFRs in aged rats. ChAT activity in these rats was lower (approximately 30%) than in adult rats in all the brain regions examined. NGF levels were not modified in cortex and hippocampus and were decreased in neostriatum (55%) and septum (35%). In conclusion, our results suggest that, during the aging process, the cholinergic impairment is related to a decrease in NGF levels in neostriatum but not in cortex and hippocampus. The reduction in level of NGF protein in septum could be due to a decrease in number of high-affinity 125I-NGF binding sites.