Turnover Rate of Brain Acetylcholine Using HPLC Separation of the Transmitter

A simple, reliable method was developed for measuring brain acetylcholine (ACh) turnover using HPLC methodology. Mice were injected intravenously with [3H]choline ([3H]Ch), and the turnover rate of ACh was calculated from the formation of [3H]ACh. Ch and ACh were separated from phosphorylcholine and from other radioactive compounds using tetraphenylboron extraction and counterion/reverse-phase chromatography. Endogenous Ch and ACh were quantified electrochemically through hydrogen peroxide production in a postcolumn reactor containing covalently bonded ACh esterase and Ch oxidase. Labeled Ch and ACh were quantified in the same sample by collecting the chromatographic fractions for radioactive content determinations. The method is rapid, well adapted to large series, and highly reproducible, with recoveries of 72.1% for Ch and 79.3% for ACh. The turnover value in mouse cerebral hemispheres was 16.02 nmol g-1 min-1 and decreased to 9.94 nmol g-1 min-1 in mice treated with oxotremorine.     

Ganglioside Composition of Normal and Mutant Mouse Embryos

The enrichment of gangliosides in neuronal membranes suggests that they play an important role in CNS development. We recently found a marked tetrasialoganglioside deficiency in twl/twl mutant mouse embryos at embryonic day (E)-11. The recessive twl/twl mutants die at embryonic ages E-9 to E-18 from failed neural differentiation in the ventral portion of the neural tube. In the present study, we examined the composition and distribution of gangliosides in twl/twl mutant mouse embryos at E-12. The total ganglioside sialic acid concentration was significantly lower in the mutants than in normal (+/-) embryos. The mutants also expressed significant deficiencies of gangliosides in the "b" metabolic pathway (GD3, GD1b, GT1b, and GQ1b) and elevations in levels of gangliosides in the "a" metabolic pathway (GM3, GM2, GM1, and GD1a). These findings suggest that the mutants have a partial deficiency in the activity of a specific sialyltransferase in the b pathway. Regional ganglioside distribution was also studied in E-12 normal mouse embryos. The ganglioside composition in heads and bodies was similar to each other and to whole embryos. Total ganglioside concentration and the distribution of b pathway gangliosides were significantly higher in neural tube regions than in nonneural tube regions. These findings suggest that b pathway gangliosides accumulate in differentiating neural cells and that the deficiency of these gangliosides in the twl/twl mutants is closely associated with failed neural differentiation.     

Acetylcholinesterase in Mouse Neuroblastoma NB2A Cells: Analysis of Production, Secretion, and Molecular Forms

The mouse neuroblastoma cell line NB2A produces cellular and secreted acetylcholinesterase (AChE). After incubation of the cells for 4 days the ratio between AChE secreted into the medium and AChE in the cells was 1:1. The cell-associated enzyme could be subdivided into soluble AChE (25%) and detergent-soluble AChE (75%). Both extracts contained predominantly monomeric AChE (4.6S) and minor amounts of tetrameric AChE (10.6S), whereas the secreted AChE in the culture supernatant contained only the tetrameric form. All forms were partially purified by affinity chromatography. It could be demonstrated that the secretory and the intracellular soluble tetramers were hydrophilic, whereas the detergent-soluble tetramer was an amphiphilic protein. On the other hand the soluble and the detergent-soluble monomeric forms were amphiphilic and their activity depended on the presence of detergent. By digestion with proteinase K amphiphilic monomeric and tetrameric AChE could be converted to a hydrophilic form that no longer required detergent for catalytic activity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of [3H]diisopropylfluorophosphate-labelled AChE gave one band at 64 kilodaltons (kD) under reducing conditions and two additional bands at 120 kD and 140 kD under nonreducing conditions.     

小鼠体内脾脏,骨髓及精原细胞姐妹染色单体交换的比较研究

本文对几种化学诱变剂诱发小鼠体内脾脏、骨髓和精原细胞的SCE进行了比较研究,同时分析了几类常见化合物在小鼠脾脏细胞中诱发SCE的活力。结果显示诱变剂在脾脏细胞中诱发SCE比骨髓和精原细胞敏感。几类化合物都能显著地诱发小鼠脾脏SCE的增加,与对照相比差异显著(P<0.05)或极显著(P<0.01),说明利用小鼠脾脏细胞检测环境诱变物是相当灵敏的。     

辽宁省番茄花叶病主要病毒和株系的研究

从辽宁南部及沈阳地区采集番茄花叶病样本111个(1985-1987),经鉴定得知主要毒原为番茄的烟草花叶病毒(TMV)占总调查的59.46％。黄爪花叶病毒(CMV)亦有发生仅占总调查的9.00％。还有一些系混合侵染(TMV和CMV)占21.62％。尚未认清毒原种的约占2.70％,接种无症的占7.20％。通过试验认为我省番茄上主要毒原为烟草花叶病毒,并进行了常规鉴定、提纯、电镜观察、光谱测定和血清测定等。对番茄上烟草花叶病毒16个分离物做了株系分化研究,按照Pelbam(1970,1972)〔14、15、16、17〕的抗性分类法应用番茄GCR一套品种接种观察,得知辽南及沈阳地区分布很广的番茄TMV分离物,均属TMV-O株系。故在番茄抗TMV育种中尚应考虑到抗1及2株系的能力。     

Changes in protein phosphorylation associated with compaction of the mouse preimplantation embryo

In order to investigate the role of protein phosphorylation in the early differentiative events of mouse preimplantation development, timed groups of embryos of various stages were incubated in medium containing [32P]orthophosphate and harvested immediately after labelling or following a chase period. The phosphoproteins obtained were separated by electrophoresis in one and two dimensions. While some of the phosphoproteins found were common to all the stages examined, the detection of many depended on both the combination of pulse-labelling and chase periods used and on the developmental stage examined. Some phosphoproteins were only found in compacted 8-cell embryos, a correlation which suggests a possible link with the post-translational mechanisms which underlie compaction.     

Influence of dietary nicotine and colony source ofManduca sexta [Lepidoptera: Sphingidae] on its suitability as a host ofCotesia congregata [Hymenoptera: Braconidae]

Larval tobacco hornworms,Manduca sexta (L.), of 2 different colonies were exposed to parasitism by the gregarious endoparasitoid,Cotesia congregata (Say). A comparison was made of parasitoid larval, pre-pupal, and pupal mortality, female and male dry weight and larval development time. In general, “Maryland” hornworms were more suitable hosts than “North Carolina” hornworms. Although the presence of dietary nicotine increased parasitoid mortality in individuals reared from hornworms of both colonies, the effect was more severe among individuals parasitizing the North Carolina hornworms. Scientific contribution No. 8125, article No. A-5066 of the Maryland Agricultural Experiment Station, Department of Entomology.     

小鼠早期胚胎发育期间TGF-β_1免疫组织化学定位

用兔抗人血小板TGF-β_1 N末端1—29氨基酸残基人工合成多肽抗血清作探针以及免疫荧光和免疫酶染色技术,分析了1—12天小鼠早期发育期间胚胎的TGF-β_1物质分布。结果表明,着床前胚胎包括卵裂细胞,桑椹胚和胚泡的ICM及滋养外胚层等细胞均显示TGF-β_1阳性免疫荧光染色。免疫酶染色还证明,沿囊胚腔顶部单层排列的原始内胚层细胞比邻近的ICM细胞有较深的染色反应。随着胚胎着床和进一步发育,7天龄胚胎中胚层早期形成阶段,紧靠中胚层一侧的外胚层胞质中含有浓集的棕色颗粒;各胚层的部分区域也存在着染色强度上差别。8—12天龄胚胎中,体节,心壁、间质细胞和肠道以及卵黄囊的脏壁中胚层均有显著的TGF-β_1免疫酶阳性物质。这些结果表明,着床前小鼠胚胎富含TGF-β_1物质,着床后的胚外组织,例如卵黄囊也为胚胎进一步发育提供了富含TGF-β_1物质的微环境;同时也提示,小鼠早期胚胎发育期间的胚泡形成,ICM细胞分化出原始内胚层,卵柱期中胚层形成,以及以后的神经管、体节和肢芽形成阶段等一系列形态发生和器官形成过程中,TGF-β_1可能是参与重要作用的一种生长调节因子。     

Control of the Amiloride-Sensitive Na+ Current in Salivary Duct Cells by Extracellular Sodium

We have previously reported that intralobular salivary duct cells contain an amiloride-sensitive Na⁺ conductance (probably located in the apical membranes). Since the amiloride-sensitive Na⁺ conductances in other tight epithelia have been reported to be controlled by extracellular (luminal) Na⁺, we decided to use whole-cell patch clamp techniques to investigate whether the Na⁺ conductance in salivary duct cells is also regulated by extracellular Na⁺. Using Na⁺-free pipette solutions, we observed that the whole-cell Na⁺ conductance increased when the extracellular Na⁺ was increased, whereas the whole-cell Na⁺ permeability, as defined in the Goldman equation, decreased. The dependency of the whole-cell Na⁺ conductance on extracellular Na⁺ could be described by the Michaelis-Menten equation with a K _m of 47.3 mmol/1 and a maximum conductance (G _max) of 2.18 nS. To investigate whether this saturation of the Na⁺ conductance with increasing extracellular Na⁺ was due to a reduction in channel activity or to saturation of the single-channel current, we used fluctuation analysis of the noise generated during the onset of blockade of the Na⁺ current with 200 μmol/l 6-chloro-3,5-diaminopyrazine-2-carboxamide. Using this technique, we estimated the single channel conductance to be 4 pS when the channel was bathed symmetrically in 150 mmol/l Na⁺ solutions. We found that Na⁺ channel activity, defined as the open probability multiplied by the number of available channels, did not alter with increasing extracellular Na⁺. On the other hand, the single-channel current saturated with increasing extracellular Na⁺ and, consequently, whole-cell Na⁺ permeability declined. In other words, the decline in Na⁺ permeability in salivary duct cells with increasing extracellular Na⁺ concentration is due simply to saturation of the single-channel Na⁺ conductance rather than to inactivation of channel activity. Received: 27 July 1995/Revised: 7 December 1995     

马桑菌株的生物学特性

对从湖北、四川、云南等地采集的尼泊尔马桑(Coriaria nepalensis Wall.)根瘤中分离的20株内生菌纯培养物所进行的研究表明,其形态均具有弗兰克氏菌属的特征。在粗细不一的分枝菌丝体上有泡囊和孢囊,有的还有串珠状结构的菌丝体。不同菌株在液体培养基上生长速度很不一致;菌丝体多呈絮状或颗粒状沉淀,个别还呈薄膜状沉淀;绝大多数菌株的菌丝体具有不同色调的色素;在S培养基中大多数都能产生可溶性色素。上述这些培养特征常因培养基成分和培养方式的不同而有差异。大多数菌株的胞壁组分属Ⅱ型,少数菌株为Ⅲ型;全细胞糖型变化很大,20个菌株可划分为6个糖型,而且绝大多数糖型与已知弗兰克氏菌有所不同。此外,有65％的菌株还具有明显的抗菌活性。