Improving effects of huperzine A on spatial working memory in aged monkeys and young adult monkeys with experimental cognitive impairment

Our previous studies demonstrated that huperzine A, a reversible and selective acetylcholinesterase inhibitor, exerts beneficial effects on memory deficits in various rodent models of amnesia. To extend the antiamnesic action of huperzine A to nonhuman primates, huperzine A was evaluated for its ability to reverse the deficits in spatial memory produced by scopolamine in young adult monkeys or those that are naturally occurring in aged monkeys using a delayed-response task. Scopolamine, a muscarinic receptor antagonist, dose dependently impaired performance with the highest dose (0.03 mg/kg, i.m.) producing a significant reduction in choice accuracy in young adult monkeys. The delayed performance changed from an average of 26.8/30 trials correct on saline control to an average of 20.2/30 trials correct after scopolamine administration. Huperzine A (0.01-0.1 mg/kg, i.m.) significantly reversed deficits induced by scopolamine in young adult monkeys on a delayed-response task; performance after an optimal dose (0.1 mg/kg) averaged 25.0/30 correct. In four aged monkeys, huperzine A (0.001-0.01 mg/kg, i.m.) significantly increased choice accuracy from 20.5/30 on saline control to 25.2/30 at the optimal dose (0.001 mg/kg for two monkeys and 0.01 mg/kg for the other two monkeys). The beneficial effects of huperzine A on delayed-response performance were long lasting; monkeys remained improved for about 24 h after a single injection of huperzine A. This study extended the findings that huperzine A improves the mnemonic performance requiring working memory in monkeys, and suggests that huperzine A may be a promising agent for clinical therapy of cognitive impairments in patients with Alzheimer's disease.

Sleeping sites of black-and-white snub-nosed monkeys (Rhinopithecus bieti) at Baima Snow Mountain, China

Data on sleeping sites of a group of black-and-white snub-nosed monkeys Rhinopithecus bieti (Colobinae, Primates) were collected between April-July and September-December 2001 to try to determine the factors affecting site selection at Nanren (99 degrees

Diet and feeding behavior of Rhinopithecus bieti at Xiaochangdu, Tibet: Adaptations to a marginal environment

The diet and feeding ecology of a wild subpopulation of black-and-white snub-nosed monkeys (Rhinopithecus bieti) were studied at Xiaochangdu in Honglaxueshan Nature Reserve, Tibet. This region is climatologically harsher than any other inhabited by non-human primates. Black-and-white snub-nosed monkeys fed on 48 parts of 25 plant species, at least three species of lichens and seven species of invertebrates. The number of food items exploited varied markedly among seasons, with dietary diversity being greatest in spring and summer. In winter, black-and-white snub-nosed monkeys had to subsist on fallback foods such as dried grass and bark. Ubiquitous lichens formed a major dietary constituent throughout the year, contributing about 75% of feeding records. Even though lichens act as a staple, our findings signify that the monkeys at Xiaochangdu prefer feeding on foliage, which is higher in protein content than the former. We provide evidence that black-and-white snub-nosed monkeys are able to cope with an array of food items other than lichens and hence can be regarded as feeding generalists. We discuss the results with reference to previous studies on other subpopulations living in habitats that are floristically more diverse and offer more plant food items than the marginal habitat at Xiaochangdu.

First description of the male and redescription of the female of Philometra clavaeceps Dogiel and Akhmerov, 1959 (Nematoda : Philometridae), a parasite of east Asian cyprinid fish

The first description of the male and a redescription of the female of the nematode Philometra clavaeceps Dogiel and Akhmerov, 1959, a parasite of east Asian cyprinids, are presented on the basis of specimens collected from Culter erythropterus Basilewsky and Culler dabryi Bleeker from Liangzi Lake (the Yangtze River basin), Hubei Province, central China. Gravid females from the fish abdominal cavity, penetrating often into ovaries, occurred in May-June, whereas conspecific males and young mature females on the swimbladder were recorded in January. Philometra clavaeceps seems to have a pronounced annual maturation cycle in the locality. The finding of P. clavaeceps in C. dabryi represents a new host record.

Flying lemurs – The 'flying tree shrews'? Molecular cytogenetic evidence for a Scandentia-Dermoptera sister clade

Background: Flying lemurs or Colugos (order Dermoptera) represent an ancient mammalian lineage that contains only two extant species. Although molecular evidence strongly supports that the orders Dermoptera, Scandentia, Lagomorpha, Rodentia and Primates form a superordinal clade called Supraprimates (or Euarchontoglires), the phylogenetic placement of Dermoptera within Supraprimates remains ambiguous. Results: To search for cytogenetic signatures that could help to clarify the evolutionary affinities within this superordinal group, we have established a genome-wide comparative map between human and the Malayan flying lemur (Galeopterus variegatus) by reciprocal chromosome painting using both human and G. variegatus chromosome-specific probes. The 22 human autosomal paints and the X chromosome paint defined 44 homologous segments in the G. variegatus genome. A putative inversion on GVA 11 was revealed by the hybridization patterns of human chromosome probes 16 and 19. Fifteen associations of human chromosome segments (HSA) were detected in the G. variegatus genome: HSA1/3, 1/10, 2/21, 3/ 21, 4/8, 4/18, 7/15, 7/16, 7/19, 10/16, 12/22 (twice), 14/15, 16/19 (twice). Reverse painting of G. variegatus chromosome-specific paints onto human chromosomes confirmed the above results, and defined the origin of the homologous human chromosomal segments in these associations. In total, G. variegatus paints revealed 49 homologous chromosomal segments in the HSA genome. Conclusion: Comparative analysis of our map with published maps from representative species of other placental orders, including Scandentia, Primates, Lagomorpha and Rodentia, suggests a signature rearrangement (HSA2q/21 association) that links Scandentia and Dermoptera to one sister clade. Our results thus provide new evidence for the hypothesis that Scandentia and Dermoptera have a closer phylogenetic relationship to each other than either of them has to Primates.

灵长类认知相关基因-Microcephalin和PACAP 前体基因的分子进化研究

人是生命演化的产物。然而由于具有一个超级大的大脑和高级的认知能力， 人类在自然界中又显得十分与众不同。在灵长类进化过程中，脑容量大小和认知 能力高低可以明显的分成几个等级，由高到低依次为人类、大猿和其它灵长类。 现在一般认为，人类超级大的大脑的形成是适应性选择(达尔文正选择)的结果。 但人类高级认知的起源一直还是个未解之谜。随着分子进化理论和比较基因组学 的发展，人们有可能从分子进化的角度去探讨高级认知的起源问题，并揭示其遗 传学机制。 本文从两个不同的角度入手，通过对认知相关的候选基因的分子进化研究来 探索高级认知起源的遗传学机制。1. 从表型出发，发现microcephalin 基因跟小 脑症相关。我们推测，该基因可能在人类起源过程中发生一些突变，导致脑容量 增大。2. 比较基因组学方法发现垂体腺苷酸环化酶激活肽(PACAP)前体基因在人 类支系中存在正选择。这类受正选择的神经相关基因可能在进化过程中对认知能 力的提高发挥重要作用。 Microcephalin 基因在人脑的发育过程中起重要作用。导致该基因蛋白质翻译 提前终止的突变，会引起人类小脑症，患者脑容量回复到早期原始人的大小。我 们对人群和12 个非人灵长类物种的microcephalin 基因编码区进行测序。所测的 非人灵长类物种包括大猿、小猿、旧大陆猴和新大陆猴。结果显示，microcephalin 基因在人群中具有较高的多态性。在人群中编码区有22 个多态位点，其中有15 个是错义突变位点。中性检验结果显示，microcephalin 基因的高多态性是人类群 体扩张和达尔文正选择共同作用的结果。灵长类各支系的进化分析表明， microcephalin 基因在大猿的起源过程中受到较强的正选择信号。这正好和小猿向 大猿进化过程中大脑容量急剧增大，认知能力大幅度提高的事实相符。单个氨基 酸编码位点检验显示，microcephalin 基因有五个氨基酸位点在灵长类的进化过程中存在正选择信号。这几个位点可能在灵长类进化和人类起源过程中，对脑容量 的增大和认知能力的提高起到了重要的作用。 PACAP 是一个神经系统中高表达的神经肽，与神经发生和神经信号传导有 关。该基因的氨基酸序列在脊椎动物的进化过程中非常保守，表明PACAP 前体 基因在这一进化历程中受到强烈的功能限制。然而比较序列分析显示，自从人与 黑猩猩分歧以后，人类支系中PACAP 前体基因有明显的加速进化。由于存在强 烈的正选择作用，人类支系的氨基酸替换速率是其它哺乳动物支系的至少7 倍。 PACAP 前体基因中有7 个人类特异的氨基酸改变，而这些氨基酸在从鼠类到大 猿的所有物种中都是保守的。蛋白质结构分析结果显示，人类起源过程中，PACAP 前体基因可能编码一个新的神经肽，并在人脑中具有功能。因此，在人类起源过 程中，PACAP 前体基因发生了适应性的改变，可能对人类认知能力的产生起到 了重要作用。这个新神经肽是否存在及其功能还有待于进一步功能实验的验证。 总之，本文通过分子进化分析，发现了两个在灵长类进化过程中对认知能力 的提高起关键作用的基因(Microcephalin 基因、PACAP 前体基因)。Microcephalin 基因，在1400-1800 万年前，从小猿向大猿的进化过程中，可能对脑容量的增大 和认知能力的提高发挥了重要的作用。PACAP 前体基因则在最近的几百万年内， 现代人的起源过程中，对人脑的形成以及人类高级认知能力的产生可能发挥了重 要的作用。

灵长类高级认知功能起源的遗传机制初探

大约5一7百万年前，从大猿中分出来的一支，走上了以大脑容量快速扩增为特征的演化道路，并最终进化成具有高级认知能力的物种-人。一直以来古人类学家和生物学家经过不懈努力，向我们揭示了人与灵长类其他物种的亲缘关系，并且让我们知道黑猩猩是同人类亲缘关系最近的现生物种。然而人口的爆炸增长和黑猩猩种群数目的急剧减少形成的鲜明对比，让我们迷惑于为何在进化地位上如此相近的物种，命运却截然不同。通过比较两个物种，科学家们发现问题的关键在于：人具有高度发达的大脑和相伴而生的高级认知能力。高度发达的认知能力使得人类获得了对环境强大的适应力，从而能够从众多生物中脱颖而出成为统治地球的智慧生物。通过比较黑猩猩和人基因组序列，生物学家发现人和黑猩猩DNA序列相似度接近99％。这个结果提示我们：1）只有很少数目的基因在一级结构上的变化和认知差异有关；2）认知差异可能更多是由调控相关元件变化导致的基因表达水平上的差异引起的。基于此，我们试图通过比较两个物种目前所知的在人脑中表达的基因的一级结构差异，找寻那些在人起源过程中受到较强正选择的基因。我们检测了近万个基因，结果显示只有大约1．38％的基因有明显的正选择迹象。接着我们从中选取了一个基因-MrgX2，测了全世界范围内的代表性人群以及非人灵长类中的代表性物种，结果显示MrgX2在人这一支受到了很强的适应选择作用，这一适应又很有可能和人类起源过程中感觉神经系统的适应有关。神经生物学的研究表明，大脑皮层前额叶是行使高级认知功能的主要区域。大脑在发育的过程中有一个神经网络重新搭建的过程，而这一过程和基因表达是密切相关的。基于此我们采用基因芯片的方法，通过比较称猴大脑发育各个关键时期的前额叶基因表达图谱，以期了解与大脑神经网络构建相关基因的活动情况。

microRNA 在灵长类中的进化及其对靶基因表达变异度的影响

microRNAs（miRNAs）是基因组中广泛编码的一类小RNA 基因，存在于绝大多数多细胞生物中，而且在各种生物学过程中都起着举足轻重的作用。miRNAs 在转录后水平通过与mRNAs 的3’UTRs 序列互补识别靶基因，并引起靶基因的降解或阻遏其翻译。在动物中，一个miRNA 可以调控数百个靶基因的表达。大多数miRNAs 在物种间高度保守，暗示了其功能的重要性。然而，非保守的miRNAs可能对物种特有新功能的产生有贡献。为了回答miRNAs是如何起源，如何进化的问题，我们研究了两个非保守miRNA 家族在灵长类中的进化历史。第一个miRNA 家族位于X 染色体上，在灵长类中的数目比狗或啮齿类中的多。我们比较了这一家族在灵长类主要分支－人、大猿、小猿、旧大陆猴和新大陆猴中的序列情况，发现了这一家族在灵长类中的快速进化。这种快速进化包括频繁的串联重复和碱基替换现象。此外，在人和黑猩猩中还发现了相应进化分支特有的替换，可能会导致分支特有的新miRNAs 的产生。对这一miRNA 家族在不同发育阶段恒河猴睾丸中的表达分析揭示了miRNA 表达变化和雄性性成熟之间的负相关，暗示这一家族在睾丸发育和精子成熟中可能起的调节作用。最后，我们认为，像蛋白编码基因一样，与雄性生殖功能相关的miRNAs 容易受到性选择而发生适应性进化。第二个miRNA 家族是位于19 号染色体上的一个灵长类特有的家族。通过分析和比较这一家族以及其临近区域在9 个不同灵长类物种中的序列，我们发现了 Alu 介导的这一家族的产生和扩张。序列比较表明，物种内和物种间miRNAs 的序列分歧相似；同时，在各个灵长类分支中均存在基因拷贝的获得和丢失，也存在基因的假基因化。由此表明，这一家族在灵长类中经历了典型的“生－死”进化历程，暗示这个家族的miRNA 基因在灵长类的进化中其功能可能发生了多样化，以适应不同灵长类物种在发育过程中的需要。此外，二级结构的保守性和前体miRNAs 区域的低SNP 密度都表明这一家族受到功能性约束。最后，我们进一步分析了这一家族在胎盘和胎儿大脑中的表达，揭示其对灵长类胚胎发育可能的重要性。除了研究miRNAs 在灵长类中的进化，我们还探讨了miRNAs 对基因表达变异度的影响。通过对已发表的193 例人类大脑基因表达谱的分析发现，基因在人群中的表达变异的大小和调控它的miRNA 数目呈正比，这暗示了miRNAs 对基因表达变异度的直接影响。相比于不受miRNA 调控的基因，受到两个以上 miRNA 核心区调控的基因有较高的表达变异度，不受miRNA 类型的影响。同时，我们还证明，人群中靶基因miRNA 识别序列上的变异（SNPs）会进一步导致靶基因表达变异的增加。我们的研究表明miRNAs 是影响人群中基因表达变异度的因素之一。

Genetic improvement on Chinese shrimp (Fenneropenaeus chinensis): growth and viability performance in F1 hybrids of different populations

Fenneropenaeus chinensis distributed in the Yellow Sea and Bohai Sea of China and the west coast of the Korean Peninsula. Different geographical populations represent potentially different genetic resources. To learn further the characteristics of different geographical population, crosses among two wild and three farmed populations were produced. The two wild populations were from the Yellow Sea and Bohai Sea (WYP), and the west coast of the Korean Peninsula and coast (WKN). The three farmed populations included the offspring of first generation of wild shrimp from coast in Korea (FKN), the Huang Hai (the Yellow Sea in Chinese) No.1 (HH1), and JK98. The phenotypes growth and survival rates of these populations were compared to confirm the feasibility for crossbreeding. The body length (BL), carapace length (CL), carapace width (CW), height of the second and third abdominal segment (HST), width of the second and third abdominal segment (WST), length of the first abdominal segment (LF), length of the last abdominal segment (LL), live body weight (BW), and survival rate were measured. Different combinations were statistically performed with ANOVA and Duncan's Multiple Range Test. The results show that the survival rate of JK98(a (TM) Euro)xWKN(a (TM),) was the highest, followed by WYP(a (TM) Euro)xWKN(a (TM),), FKN(a (TM) Euro)xWYP(a (TM),), FKN(a (TM) Euro)xHH1(a (TM),) and WYP(a (TM) Euro)xFKN(a (TM),); the body weight of FKN(a (TM) Euro)sxHH1(a (TM),) was the highest, followed by FKN(a (TM) Euro)xWYP(a (TM),), WYP(a (TM) Euro)xWKN(a (TM),), WYP(a (TM) Euro)xFKN(a (TM),) and JK98(a (TM) Euro)xWKN(a (TM),); the total length had the same ranking as the body weight. All growth traits in hybrids JK98(a (TM) Euro)xWKN(a (TM),) were the lowest among all combinations. F1 hybrids had significant difference (P < 0.05) in BL, CL, HST, LL, and BW; and insignificant difference (P > 0.05) in other growth traits and survival rate. The results of Duncan's Multiple Range Test are that BL and CL of JK98(a (TM) Euro)xWKN(a (TM),) were significantly different from the other combinations; HST different from the combination of FKN(a (TM) Euro)xWYP(a (TM),), FKN(a (TM) Euro)xHH1(a (TM),) and WYP(a (TM) Euro)xWKN(a (TM),); and BW different from FKN(a (TM) Euro)xWYP(a (TM),) and FKN(a (TM) Euro)xHH1(a (TM),). As a whole, the results indicate that the FKN(a (TM) Euro)xHH1(a (TM),) was the best combination in all growth traits. Therefore, hybridization can introduce the variation to base populations. The systematic selection program based on additive genetic performance may be more effective than crossbreeding.

Highly sensitive determination of the methylated p16 gene in cancer patients by microchip electrophoresis

The p16 tumor suppressor gene is inactivated by promoter region hypermethylation in many types of tumor. Recent studies showed that aberrant methylation of the p16 gene is an early event in many tumors, especially in lung cancer, and may constitute a new biomarker for early detection and monitoring of prevention trials. We detected tumor-associated aberrant hypermethylation of the p16 gene in plasma and tissue DNA from 153 specimens using a modified semi-nested methylation-specific PCR (MSP) combining plastic microchip electrophoresis or slab gel electrophoresis, respectively. Specimens were from 79 lung cancer patients, 15 abdominal tumor patients, 30 positive controls and 30 negative controls. The results showed that the positive rate obtained by microchip electrophoresis was more than 26.6% higher and the same speciticity was kept when compared with slab gel electrophoresis. The microchip electrophoresis can rapidly and accurately analyze the PCR products of methylated DNA and obviously improve the positive rate of diagnosis of cancer patients when compared with gel electrophoresis. This method with the high assay sensitivity might be used for detection of methylation of p16 gene and even to facilitate early diagnosis of cancer patients. (C) 2004 Elsevier B.V. All rights reserved.