HORMONAL-REGULATION OF TISSUE-TYPE PLASMINOGEN-ACTIVATOR AND PLASMINOGEN-ACTIVATOR INHIBITOR TYPE-1 IN CULTURED MONKEY SERTOLI CELLS

Sertoli cells play a central role in the control and maintenance of spermatogenesis. Isolated Sertoli cells of mouse and rat testes have been shown to secrete plasminogen activator (PA) and a plasminogen activator inhibitor type-1 (PAI-1) in culture. In this study, we have investigated the hormonal regulation of PA and PAI-1 activities in cultured monkey Sertoli cells. Sertoli cells (5x10(5) cells/well) isolated from infant rhesus monkey testes were preincubated at 35 degrees C for 16 h in 24-well plates precoated with poly(D-lysine) (5 mu g/cm(2)) in 0.5 mi McCoy's 5a medium containing 5% of fetal calf serum and further incubated for 48 h in 0.5 mi serum-free medium with or without various hormones or other compounds, PA as well as PAI-1 activities in the conditioned media were assayed by fibrin overlay and reverse fibrin autography techniques respectively. The Sertoli cells in vitro secreted only tissue-type PA (tPA), no detectable amount of urokinase-type PA (uPA) could be observed, Monkey Sertoli cells were also capable of secreting PAI-1, Immunocytochemical studies indicated that both tPA and PAI-1 positive staining localized in the Sertoli cells, spermatids and residual bodies of the seminiferous epithelium; Northern blot analysis further confirmed the presence of both tPA and PAI-1 mRNA in monkey Sertoli cells. Addition of follicle-stimulating hormone (FSH) or cyclic adenosine monophosphate (cAMP) derivatives or cAMP-generating agents and gonadotrophin-releasing hormone (GnRH) agonist or phorbol ester (PMA) to the cell culture significantly increased tPA activity. PAI-1 activity in the culture was also enhanced by these reagents except 8-bromo-dibutyryl-cAMP, forskolin and 3-isobutyl-1-methylxanthin (MIX) which greatly stimulated tPA activity, whereas decreased PAI-1 activity, implying that neutralization of PAI-1 activity by tile high level of tPA in the conditioned media may occur. These data suggest that increased intracellular signals which activate protein kinase A (PKA), or protein kinase C (PKC) can modulate Sertoli cell tPA and PAI-1 activities, The concomitant induction of PA and PAI-1 by the same reagents in the Sertoli cells may reflect a finely tuned regulatory mechanism in which PAI-1 could limit the excession of the proteolysis.

HORMONAL REPLACEMENT THERAPY IN FISH - HUMAN GROWTH-HORMONE GENE-FUNCTION IN HYPOPHYSECTOMIZED CARP

Transgenic common carp, Cyprinus carpio, produced by the microinjection of fertilized eggs with a linearized chimeric plasmid pMThGH, a human growth hormone (hGH) gene with a mouse metallothionein-I (MT) gene promoter in pBR322, were used to produce F1 and F2 transgenics. Following hypophysectomy of the transgenic F2 common carp, non-transgenic common carp and non-transgenic crucian carp, growth was monitored for up to 110 days. In addition, recombinant hGH was injected subcutaenously into a group of the non-transgenic crucian carp. Growth rate analyses indicated that (1) hypophysectomy of non-transgenic common carp and crucian carp results in the cessation of growth, (2) hGH administration can stimulate the growth of hypophysectomized crucian carp and (3) hypophysectomized hGH-transgenic common carp continue to grow in the absence of their own growth hormone, suggesting that the hGH-transgene is being expressed in tissues other than the pituitary.

Neuro-Stimulus Chip with Photodiodes Array for Sub-retinal Implants

A prototype neuro-stimulus chip for sub-retinal implants in blind patients affected by Age-related Macular Degeneration (AMD) or Retinitis Pigmentosa (RP) is presented in this paper. This retinal prosthetic chip was designed to replace the degenerated photoreceptor cells, and in order to stimulate directly the remaining healthy layers of retinal neurons. The current stimulus circuits are monolithic integrated with photodiodes (PD) array, which can convert the illumination on the eyes into bi-phasic electrical pulses. In addition, a novel charge cancellation circuit is used to discharge the electrodes for medical safty. The prototype chip is designed and fabricated in HJTC 0.18 mu m N-well CMOS 1P6M Mix-signal process, with a +/- 2.5 V dual voltage power supply.

Peptidomics and genomics analysis of novel antimicrobial peptides from the frog, Rana nigrovittata

Much attention has been paid on amphibian peptides for their wide-ranging pharmacological properties, clinical potential, and gene-encoded origin. More than 300 antimicrobial peptides (AMPs) from amphibians have been studied. Peptidomics and genomics analysis combined with functional test including microorganism killing, histamine-releasing, and mast cell degranulation was used to investigate antimicrobial peptide diversity. Thirty-four novel AMPs from skin secretions of Rana nigrovittata were identified in current work, and they belong to 9 families, including 6 novel families. Other three families are classified into rugosin, gaegurin, and temporin family of amphibian AMP, respectively. These AMPs share highly conserved preproregions including signal peptides and spacer acidic peptides, while greatly diversified on mature peptides structures. In this work, peptidomics combined with genomics analysis was confirmed to be an effective way to identify amphibian AMPs, especially novel families. Some AMPs reported here will provide leading molecules for designing novel antimicrobial agents. (C) 2009 Elsevier Inc. All rights reserved

氟代柠檬酸对体外培养的神经胶质瘤细胞G422增殖的抑制效应

为研究氟代柠檬酸(Fluorocitrate)对体外培养的神经胶质瘤细胞生长的影响,采用MTT法研究不同的氟代柠檬酸浓度(0.0025mmol/L,0.005mmol/L,0.01 mmol/L,0.025mmol/L和0.1 mmol/L)和作用时间(36h,48h和60h)对神经胶质瘤细胞G422增殖的影响.结果发现:(1)氟代柠檬酸可抑制G422细胞的增殖,并且其抑制作用随氟代柠檬睃浓度的增加而增强;(2)高浓度(0.01 mmol/L,0.025 mmol/L和0.1 mmol/L)氟代柠檬酸对G422细胞的增殖抑制作用随作用时问的延长而增强:(3)低浓度(0.0025mmol/L和0.005mmol/L)氟代柠檬酸对G422细胞的增殖抑制作用不随作用时间的延长而改变.实验表明,氟代柠檬酸能够抑制神经胶质瘤细胞的增殖,其抑制能力随氟代柠檬酸浓度的增加和作用时间的延长而加强.

Delta EEG Activity in Left Orbitofrontal Cortex in Rats Related to Food Reward and Craving

眶额叶皮质与中脑边缘多巴胺奖赏系统有着复杂的相互纤维联系.先前的研究探讨了药物成瘾过程中眶额叶皮质的脑电活动.在本实验中,将探讨食物奖赏和渴求过程中该皮质的脑电活动.实验采用了两个环境:对照环境和食物刺激相关的环境.首先,训练大鼠在食物刺激相关的环境中吃巧克力花生豆,而后在该环境中设置两种不同的刺激方式:能看到和闻到但不能吃到(渴求实验),或者仍旧可以吃到巧克力花生豆(奖赏实验):同时进行左侧眶额叶皮质的脑电记录.结果发现,在食物刺激相关的环境中大鼠Delta频段(2-4Hz)的脑电活动与食物刺激显著相关,此外,与在对照环境中相比,其相对功率在食物渴求时下降而在食物奖赏时升高.本实验表明,食物相关的奖励可以改变大鼠眶额叶皮质的脑电活动,而且,Delta频段的脑电活动能够作为监测该奖励的一个指标.

Morphine and propranolol co-administration impair consolidation of Y-maze spatial recognition memory

In the present study, the interaction between morphine and the beta-adrenergic receptor antagonist, propranolol (PROP), in memory consolidation was investigated in a two-trial recognition Y-maze task. Four sets of Y-maze experiments were carried out in mi

Distracters Impair and Create Working Memory-Related Neuronal Activity in the Prefrontal Cortex

The prefrontal cortex (PFC) has a central role in working memory (WM). Resistance to distraction is considered a fundamental feature of WM and PFC neuronal activity. However, although unexpected stimuli often disrupt our work, little is known about the un

Pentylenetetrazole-induced status epilepticus following training does not impair expression of morphine-induced conditioned place preference

Learning and memory play an important role in morphine addiction. Status epilepticus (SE) can impair the spatial and emotional learning and memory. However, little is known about the effects of SE on morphine-induced conditioned place preference (CPP). Th

Effects of testosterone undecanoate as a male contraceptive candidate on rat immunological features

Testosterone undecanoate (TU) is under phase III clinical trial as a hormonal male contraceptive in China. Sex hormones can modulate the immune system. Female hormonal contraceptives may affect SIV/HIV-1 transmission. To evaluate the safety of TU and to understand whether long-term use of TU for a male contraceptive affects users' immunological features, adult male rats were treated for a 32-week TU-treated phase at the dose of 20 mg TU/kg body weight and a 24-week recovery phase. The reproductive and immunological parameters of 4-6 rats in each subgroup were examined at the stated time point. The mean sperm count and viability in the treated rats were significantly suppressed (p < 0.01). In the TU-treated group: the mean blood leukocyte and lymphocyte counts; the proliferation indexes of T cells from peripheral blood mononuclear cells (PBMC) and spleen; and, of B cells from spleen, as well as the mean counts of blood T, NK, and B cells decreased in comparison with those of control group. These decreases were not significant (p > 0.01). Similarly, the mean serum IgM, IgG, and IgA levels and complement activity in TU-treated rats were lower than those in control group (p > 0.01), and the changes in the antibody levels of the examined genital secretions were not significant (p > 0.01). The changes in the thickness of urethra epithelium, and in secretory component (SC) expression in genitals were not observed in the treated group. These results demonstrated that long-term supraphysiological TU injection did not obviously affect the examined rat immunological parameters.

钠离子通道类毒素的细胞毒性和检测方法的研究

赤潮毒素广泛存在于各种赤潮藻和各类海洋生物中，不仅对渔业、养殖业危害甚大，而且还直接威胁着人类的生存健康。其中，离子通道类毒素是一类毒性较高的毒素。除一些赤潮藻可以产生此类毒素之外，海洋中还存在某些生物也能够产生离子通道类毒素。为进一步阐明钠离子通道类毒素对细胞的毒性效应机制，本文选取一株小鼠神经母细胞瘤（Neuro-2a）作为受试对象，研究了四种钠离子通道类毒素STX、GTX1,4、GTX2,3、TTX对Neuro-2a细胞的毒性影响机制，并利用STX和TTX，建立了钠离子通道类毒素的细胞毒性检测方法，且应用此方法检测了贝体内、藻体内的毒素含量，进一步与小鼠法和HPLC法进行了比较。 研究表明：STX、GTX1,4、GTX2,3、TTX四种钠离子通道类毒素在长时间内均会对Neuro-2a细胞的增殖产生不利影响。在短时间（24h）内，以上各毒素均没有抑制Neuro-2a细胞的增殖，但是48h后，以上各毒素对Neuro-2a细胞的增殖均产生了抑制作用，且随着各毒素剂量的增加，细胞增殖受抑制程度也表现出一定程度的增高，二者呈剂量-反应关系。STX、GTX1,4、GTX2,3、TTX对Neuro-2a细胞的48h半数抑制浓度（IC50）分别为：250ng/ml、1000ng/ml、1300ng/ml、700ng/ml。本论文还首次研究了STX、GTX1,4、GTX2,3、TTX四种钠离子通道类毒素对Neuro-2a细胞内酶活性的影响。研究发现，STX、GTX1,4、GTX2,3、TTX四种钠离子通道阻断剂类毒素均能够影响Neuro-2a细胞内Na+-K+-ATP酶和乙酰胆碱酯酶TChE的活性。当各毒素作用24h后，Neuro-2a细胞内Na+-K+-ATP酶和乙酰胆碱酯酶TChE的活力均会受到抑制，并且随着各毒素剂量的增加，两种酶的活性也逐渐降低。可见，钠离子通道阻断剂类毒素能对细胞内酶的功能产生一定的影响，此影响连同阻断细胞膜钠离子通道，造成离子流的失衡作用，进一步对细胞产生毒性效应。在对细胞膜通透性的研究中发现，上述四种钠离子通道阻断剂类毒素各剂量组细胞培养液乳酸脱氢酶LDH的漏出率与对照组相比均无显著差异，它们均未引起Neuro-2a细胞膜内LDH的改变，看来钠离子通道阻断剂类毒素不会通过影响细胞膜的通透性而对细胞引起毒性效应。 本研究还利用STX和TTX两种钠离子通道标准毒素以及乌苯苷、藜芦定两种生物毒素，参照Jellett（1992）方法，建立了STX和TTX两种钠离子通道类毒素的细胞毒性检测的标准曲线，分别为：Y=0.266X+51.184和 Y=1.6068X+47.186。检出限分别为5ng/ml和0.8ng/ml。并且利用已建立的细胞毒性检测方法检测了来自浙江舟山和连云港赣榆市的19个织纹螺样品和5株实验室培养的亚历山大藻，得到的实验结果与小鼠生物测试和HPLC检测的结果存在较好的相关关系。鉴于该方法具有高通量、省时、检出限低等优点，因此更具有在沿海环境检测中推广应用的潜力。

DSP等赤潮藻毒素对哺乳类细胞的毒性效应及机制研究

本文研究了腹泻性贝毒的主要组分大田软海绵酸以及几株重要赤潮藻的提取物对四株哺乳类细胞的毒性效应，并探讨了应用细胞毒性检测方法作为赤潮藻毒素毒性检测常规方法的可能性。 结果发现：OA和利玛原甲藻提取物显著地抑制四株细胞的增殖并诱导四株细胞发生凋亡；四株细胞对毒素的敏感性存在一定的差异，人肝癌细胞和小鼠神经瘤细胞较敏感，其次分别为人肝细胞和小鼠皮肤细胞。 小鼠神经瘤细胞对OA反应敏感，细胞毒性检测指标及检测方法灵敏、快速，应用小鼠神经瘤细胞Neuro-2a进行的DSP毒素细胞毒性测试方法具有发展为该类毒素毒性监测常规方法的潜能。 米氏凯伦藻内存在抑制细胞增殖的毒性物质，且该物质是一种具有一定极性的脂溶性物质；该物质能够导致细胞肿胀、破裂，并诱导细胞发生脂质过氧化，导致脂质过氧化产物丙二醛(MDA)的积累。 相关亚历山大藻的去藻过滤液内存在抑制细胞增殖和诱导细胞凋亡的毒性物质，该毒性物质的分子量＞5K，这与本实验室以往的研究结果一致。 　　总之，通过我们的研究发现：DSP等赤潮藻毒素或毒性物质对哺乳类细胞存在毒性影响，且不同毒素的危害机制存在差异，赤潮藻毒素或赤潮藻产生的一些毒性物质威胁人类的健康，应引起我们的高度关注。

去甲肾上腺素刺激高原鼠兔CRF的分泌

We studied the effect of neuro transmitter no repin ephrine (N E) on immuno reactive cortico trop inreleasing factor (CRF) of median eminence (M E) in the native pika (Ochotona cu rz oniae). At one hour after intra cerebrovent ricular ( icv) adm inistrat ion of N E in doses of 3.75,7.5, 15 and 30 μg/100 g BW , the CRF level ofM E increased. And the plasma cortico sterone concent rat ion also increased. Two and six days after adrenalectomy (ADX) , N E concent ration in hypothalamus declined to 76.32% and 76.27% of those in intact pika, plasma cortico rsterone concent ration also decreased to 16.57 and 2.05% of the control. These results indicated that N E have a effect on activating HPA axis through activating hypothalamic CRF in Ochotona curzoniae.

Seasonal thermogenesis and body mass regulation in plateau pikas (Ochotona curzoniae)

Changes in photoperiod, ambient temperature and food availability trigger seasonal acclimatization in physiology and behavior of many animals. In the present study, seasonal adjustments in body mass and in several physiological, hormonal, and biochemical markers were examined in wild-captured plateau pikas (Ochotona curzoniae) from the Qinghai-Tibetan plateau. Our results showed that plateau pikas maintained a relatively constant body mass throughout the year and showed no seasonal changes in body fat mass and circulating levels of serum leptin. However, nonshivering thermogenesis, cytochrome c oxidase activity, and mitochondrial uncoupling protein 1 (UCP1) contents in brown adipose tissues were significantly enhanced in winter. Further, serum leptin levels were positively correlated with body mass and body fat mass while negatively correlated with UCP1 contents. Together, these data suggest that plateau pikas mainly depend on increasing thermogenic capacities, rather than decreasing body mass, to cope with cold, and leptin may play a potential role in their thermogenesis and body mass regulation.

去甲肾上腺素参与吗啡神经适应性改变的中枢机制

That relapse still exists even after prolonged withdrawal is a difficult issue in the medical cure of drug addiction. Neuro-adaptation induced by prolonged exposure to addictive drugs is the neural mechanisms of both compulsive drug seeking and relapse.Neuro-adaptation caused by addictive drugs increases the individuals’ response to drugs and on the other hand, it reduces the response to natural reward in withdrawn individuals.There must be common neural mechanisms between the co-existing phenomena, and there must also be unique neural mechanisms in the drugs.To reveal the neuro-adaptation arising in the process from random, controllable drug-use to uncontrollable compulsive drug seeking is of great significance both theoretically and practically.Based on the above hypothesis, in order to reveal the function of alpha adrenergic receptor in compulsive drug-seeking motivation during the process of drug addiction, using sensitization of morphine-induced psychomotor activity as behavioral model, through the method of behavioral pharmacology, the neural mechanisms of alpha adrenergic receptor’s involvement in the process of addiction has been studied.The adjustment function caused by alpha receptors in medial prefrontal cortex and nucleus accumbens to morphine-induced psychomotor activity has been compared in the period of first use of drugs and in repetitive-use period. Furthermore, the effect on novelty seeking caused by alpha-receptors in relevant brain areas has also been compared. Major results are as follow: 1 After prolonged morphine exposure, rats’ response to morphine-induced psychomotor activity is strengthened and response to novel object induced reward weakened. 2 Injection of prazosin in medial prefrontal cortex will block morphine-induced psychomotor activity of naïve rats, however, it will not block that of morphine-withdrawn rats, but it will block the novelty seeking behavior of morphine-withdrawn rats. 3 Injection of clonidine in medial prefrontal cortex will block morphine-induced psychomotor effect of both naïve rats and morphine-withdrawn rats, and will block the novelty seeking behavior of morphine-withdrawn rats. 4 Injection of prazosin in nucleus accumbens will not affect the morphine-induced psychomotor effect of either naïve rats or morphine-withdrawn rats, nor will it affect the novelty seeking behavior of morphine-withdrawn rats. 5 Injection of clonidine in nucleus accumbens will block morphine-induced psychomotor effect of naïve rats, however, it will not block that of morphine-withdrawn rats, nor will it affect the novelty seeking behavior of morphine-withdrawn rats. These results show: 1 The weakening of the function of alpha1 receptors in medial prefrontal cortex and alpha2 receptors in nucleus accumbens caused by repetitive exposure to morphine is probably the cause of compulsive drug-seeking activity. 2 Blocking alpha1 receptors in medial prefrontal cortex accelerates the loss of interest in natural reward after morphine withdrawal. 3 Blocking alpha2 receptors in medial prefrontal cortex not only restrains drug-seeking motivation, but also blocks the individual’s seeking motivation for novelty stimulus, which suggests that, while selecting medicine for curing addiction, it should be considered to reduce the influence on natural reward as much as possible and to avoid major side-effect.