K+ channel cAMP activated in guinea pig gallbladder epithelial cells

In guinea pig gallbladder epithelial cells, an increase in intracellular cAMP levels elicits the rise of anion channel activity. We investigated by patch-clamp techniques whether K(+) channels were also activated. In a cell-attached configuration and in the presence of theophylline and forskolin or 8-Br-cAMP in the cellular incubation bath, an increase of the open probability (P(o)) values for Ca(2+)-activated K(+) channels with a single-channel conductance of about 160 pS, for inward current, was observed. The increase in P(o) of these channels was also seen in an inside-out configuration and in the presence of PKA, ATP, and cAMP, but not with cAMP alone; phosphorylation did not influence single-channel conductance. In the inside-out configuration, the opioid loperamide (10(-5) M) was able to reduce P(o) when it was present either in the microelectrode filling solution or on the cytoplasmic side. Detection in the epithelial cells by RT-PCR of the mRNA corresponding to the alpha subunit of large-conductance Ca(2+)-activated K(+) channels (BK(Ca)) indicates that this gallbladder channel could belong to the BK family. Immunohistochemistry experiments confirm that these cells express the BK alpha subunit, which is located on the apical membrane. Other K(+) channels with lower conductance (40 pS) were not activated either by 8-Br-cAMP (cell-attached) or by PKA + ATP + cAMP (inside-out). These channels were insensitive to TEA(+) and loperamide. The data demonstrate that under conditions that induce secretion, phosphorylation activates anion channels as well as Ca(2+)-dependent, loperamide-sensitive K(+) channels present on the apical membrane.     

Clonal propagation,encapsulation and reintroduction of <Emphasis Type="Italic">Ipsea malabarica</Emphasis> (Reichb. f.) J. D. Hook., an endangered orchid

Summary Rapid clonal propagation and encapsulation of in vitro-formed bulbs of Ipsea malabarica (Reichb. f.) J. D. Hook., an endemic and endangered orchid of the Western Ghats of Kerala, and its reintroduction to the natural habitat were accomplished. Rhizome segments of Ipsea cultured on half-strength Murashige and Skoog (MS) medium supplemented with 6.97 μM kinetin induced the highest number of shoots, at the rate of four shoots per explant within 50 d. Transfer of the isolated shoots increased the rate of shoot multiplication to more than 10 shoots. Subsequent culture enhanced the number of shoots. No decline of shoot multiplication was observed up to the 10th subculture. Shoots developed bulbs during culture which developed into rhizomes. Sucrose at 6–8% reduced the time for the development of bulbs and rhizomes. Roots were developed from the base of the developed shoots as well as from the bulbs. Isolation and culture of bulbs also developed 5–10 shoots within 50 d. Encapsulated in vitro-formed bulbs cultured either on hormone-free halfstrength MS or 6.97 μM kinetin-supplemented medium facilitated 100% conversion. As a step to conservation in situ, 50 plantlets were reintroduced into their natural habitat, i.e. at Vellarimala (at 1300 m height) of the Western Ghats of Kerala, and flowered normally. Development of more than 40 000 plantlets starting from a single explant is possible within 250 d. This threatened endemic orchid stands to benefit greatly from the established protocol and will hopefully curtail the threat of extinction.     

The use of floral homeotic mutants as a novel way to obtain durable resistance to insect pests

We have developed a novel strategy for the introduction of durable insect resistance in crops. This strategy was based on intervention in the natural relationship between plants and insects. For many insects, including pests such as thrips (Frankliniella occidentalis), the flower is an important factor in their life cycle, serving either as a food source or as a place for mating. The insects are attracted to the flower by scent, which is mainly produced by the petals, and by the bright colour of these floral organs. We therefore anticipated that removal or changing the identity of the petals would significantly reduce the attractiveness of the flower to thrips. To test this hypothesis, we used cucumber as a model species because most modern varieties are parthenocarpic, in which the fruit develops without fertilization. The cucumber mutant green petals, in which the petals are homeotically transformed into green sepals, was particularly useful for this study. The susceptibility of the cucumber plants to damage by thrips was determined by recording thrip numbers and by measuring leaf damage. Large differences were observed when greenhouse compartments with either wild-type or green petal mutant plants were compared. The rate of population growth of the insects on the mutant plants was significantly reduced and the leaves were almost undamaged. These results demonstrate that alterations in the structure of flowers may interfere with the life cycle of insects, providing the means for a novel and natural strategy for obtaining insect resistance.     

缺血引起沙土鼠纹状体DARPP—32磷酸化状态的变化

采用蒙古沙土双侧颈总动脉阻断前脑缺血模型,以放射自显影（反向磷酸化,back-phosphorylation)及免疫印迹（Western blotting)法体外测定缺血时纹状体DARPP－32磷酸化水平和蛋白含量的变化,结果表明,短暂性缺血纹状体DARPP－32的免疫学活性和蛋白含量地明显改变。在缺血10min内,随缺血时间的延长,体外DARPP－32的[^32P]的掺入量在缺血5min时升高,在缺血2,7,10min时均降低,而反向磷酸化的测定结果表明体内DARPP－32磷酸化水平增高,说明缺血可诱导DARPP－32磷酸化水平变化。     

Decreased protein levels of nicotinic receptor subunits in the hippocampus and temporal cortex of patients with Alzheimer's disease

Deficits of cortical nicotinic acetylcholine receptors (nAChRs) have been observed in Alzheimer's disease (AD) by receptor binding assays. Little is known about the receptor subunit specificity influenced by AD, and it might be of importance for therapeutic strategies. In the present study, the protein levels of nAChR alpha3, alpha4, alpha7, and beta2 subunits were investigated using western blot analysis on postmortem brains of patients with AD and age-matched controls. The results showed that in human postmortem brain samples, bands with molecular masses of 52, 42, and 50 kDa were detected by anti-alpha4, anti-alpha7, and anti-beta2 antibodies, respectively. When anti-alpha3 antibody was used, one major band of 49 kDa and two minor bands of 70 and 38 kDa were detected. In AD patients, as compared with age-matched controls, the alpha4 subunit was reduced significantly by approximately 35 and 47% in the hippocampus and temporal cortex, respectively. A significant reduction of 25% in the alpha3 subunit was also observed in the hippocampus and a 29% reduction in the temporal cortex. For the alpha7 subunit, the protein level was reduced significantly by 36% in the hippocampus of AD patients, but no significant change was detected in the temporal cortex. In neither the hippocampus nor the temporal cortex was a significant difference observed in the beta2 subunit between AD patients and controls. These results reveal brain region-specific changes in the protein levels of the nAChR alpha3, alpha4, and alpha7 subunits in AD.     

豌豆中可与Gro EL抗体发生免疫学反应的热激诱导蛋白

通过Ｗｅｓｔｅｒｎ印渍方法,发现在豌豆（ＰｉｓｕｍｓａｔｉｖｕｍＬ．）叶存在一种高分子量的蛋白可与ＧｒｏＥＬ抗体发生免疫学反应,它含有两种亚基,分子量分别为６０．４ｋＤ（α）和６５．５ｋＤ（β）,具有弱的ＡＴＰａｓｅ活性。热激处理后此蛋白的表达升高３～４倍,但外源ＡＢＡ处理并不增加它的表达。用３５Ｓ标记蛋白和氯霉素抑制实验表明,此蛋白可能是在８０Ｓ核糖体上合成的胞质蛋白。     

Reversible decreases in ATP and PCr concentrations in anoxic turtle brain

A hallmark of anoxia tolerance in western painted turtles is relative constancy of tissue adenylate concentrations during periods of oxygen limitation. During anoxia heart and brain intracellular compartments become more acidic and cellular energy demands are met by anaerobic glycolysis. Because changes in adenylates and pH during anoxic stress could represent important signals triggering metabolic and ion channel down-regulation we measured PCr, ATP and intracellular pH in turtle brain sheets throughout a 3-h anoxic-re-oxygenation transition with ³¹P NMR. Within 30 min of anoxia, PCr levels decrease 40% and remain at this level during anoxia. A different profile is observed for ATP, with a statistically significant decrease of 23% occurring gradually during 110 min of anoxic perfusion. Intracellular pH decreases significantly with the onset of anoxia, from 7.2 to 6.6 within 50 min. Upon re-oxygenation PCr, ATP and intracellular pH recover to pre-anoxic levels within 60 min. This is the first demonstration of a sustained reversible decrease in ATP levels with anoxia in turtle brain. The observed changes in pH and adenylates, and a probable concomitant increase in adenosine, may represent important metabolic signals during anoxia.     

小麦丛矮病毒M蛋白基因的序列测定,表达和鉴定

从小麦丛矮病毒（ＷＲＳＶ）基因文库含磷蛋白ＮＳ基因的质粒中,经亚克隆、测序,得到ＮＳ蛋白基因的下游序列,其中含有一读框４５３ｎｔ、编码一分子量约１７ｋＤ蛋白。用ＰＣＲ方法获得该读框全长片段并克隆到表达载体ｐＧＥＸ－３Ｘ,在大肠杆菌ＢＬ２１（ＤＥ３）菌株中以ＩＰＴＧ诱导表达,产物由谷胱甘肽Ｓ－转移酶（ＧＳＴ）亲和层析柱纯化后,用蛋白质印迹分析鉴定,结果表明,该读框为编码病毒基质蛋白Ｍ的基因。     

东喜马拉雅地区雀儿豆属的修订

本文依据形态特征对分布于东喜马拉雅地区的雀区豆属植物进行了研究,认为荚果圆柱形,长圆形并且果皮具海绵状结构非该类植物所特有;垫状生活型则是植物对高山带,荒砾质带生态适应的结果。