洋葱内表皮细胞中的网状结构物

使用 Triton 抽提和卡马氏亮蓝染色方法,在洋葱鳞茎内表皮细胞中观察到网状结构物的存在。秋水仙素和细胞松弛素 B 的处理,进一步区分出两种不同的网状结构:近质膜较细的网状结构能被秋水仙素破坏;核周围与核成切线状结合的较粗的网状结构,经细胞松弛素 B 处理后,其形象更为突出。本文讨论了卡马氏亮蓝方法所揭示的带壁细胞的网状结构物。     

The Effect of Colchicine on Regenerating Membranellar Cilia in Stentor coeruleus

SYNOPSIS. Stentors treated with toxic substances can be induced to shed their oral bands (19, Fig. 1), complex structures composed of many cilia organized into membranelles. Regenerating membranellar bands were observed in control stentors removed from toxic (urea-containing) medium at about 3.5 hours. At 8 hours regenerated control organisms were indistinguishable from normal unshed stentors. Experimental animals replaced into colchicine medium were inhibited from regeneration at low, nontoxic concentrations of this mitotic spindle inhibitor. Upon removal of the colchicine and replacement of the shed animals into normal medium or normal medium to which GTP had been added, complete and normal regeneration of the membranellar band ensued. Our observations are consistent with many suggesting that colchicine acts by reversibly binding with a protein during processes involving microtubule formation. Colchicine inhibition of membranellar band formation further indicates that oral membranelles are specialized evolutionary homologs to other centriole (= basal body, = kinetosome) derivatives such as mitotic spindle fibers, cilia and flagella, axopods, etc. (structures containing the ubiquitous microtubules of eukaryotic cells).     

Doubled haploid plants following colchicine treatment of microspore-derived embryos of oilseed rape (<Emphasis Type="Italic">Brassica napus L.</Emphasis>)

An efficient method for producing doubled haploid plants of oilseed rape (Brassica napus L.) was established using in vitro colchicine treatment of haploid embryos. Haploid embryos in the cotyledonary stage were treated with one of four colchicine concentrations (125, 250, 500 and 1,000 mg/L); for one of three treatment durations (12, 24 and 36 h) at one of the two temperatures (8 and 25°C) and were compared to control embryos (without colchicine treatment). The number of chromosomes, seed recovery, size and density of leaf stomata, and pollen grain size from regenerated plants were determined. No doubled haploid plants were regenerated from control embryos; however, the doubled haploid plants were regenerated from colchicine-treated embryos. A high doubling efficiency, 64.29 and 66.66% of regenerated plants, was obtained from 250 mg/L colchicine treatment for 24 h and 500 mg/L colchicine treatment for 36 h, respectively, at 8°C. Following 500 mg/L colchicine treatment for 36 h, a few plants regenerated (9 plants). At the higher colchicine concentration (1,000 mg/L), no plant regenerated. These results indicate that the colchicine treatment of embryos derived from microspores can induce efficient chromosome doubling for the production of doubled haploid lines of oilseed rape.     

接种叶锈菌的小麦叶片胞间洗脱液对叶肉细胞原生质体微丝骨架的影响

用异硫氰酸 鬼笔环肽 (FITC Ph)标记和共聚焦激光扫描显微镜 (CLSM )观察发现 ,以具有激发子活性的接种叶锈菌的小麦叶片的胞间洗脱液 (IWF)处理叶肉细胞原生质体一定时间后 ,抗病小麦品种洛夫林 10的原生质体内微丝骨架保持完整的网络状结构 ,而感病品种 5 389的大部分微丝骨架处于解聚状态。同时 ,抗病品种因IWF处理诱发的防卫反应———H2 O2 突发和HR反应的程度 (用原生质体活力下降的程度表示 )也大大高于感病品种。用细胞松弛素D(CD)预处理抗病品种原生质体可以明显抑制IWF处理诱发的H2 O2突发和HR反应 ,表明微丝骨架的状态可能与抗病性有密切的关系 ,完整的微丝骨架是H2 O2 突发和HR反应的一个重要条件。     

Effect of cell cycle synchronization on the accuracy of murine and bovine embryo sex determination

Different cell cycle synchronization methods were used to increase the mitotic index and accuracy of sex determination in murine and bovine embryos. For sexing purposes, colchicine treatment for 2, 4, 6 and 8 h and the FdU-thymidine-colchicine combination were tested in murine embryos. The best results were obtained with colchicine treatment for 8 h (96.88% accuracy) and with FdU-thymidine-colchicine (97.22% accuracy). Mitotic indexes differed significantly between the 2 treatments (21.71% for colchicine and 32.95% for FdU-thymidine-colchicine). For sex identification of murine and bovine demi-embryos, both treatments were demonstrated to be equally effective (nearly 90%). The mitotic index for the FdU-treated murine demi-embryos (19.04%) was higher than the one obtained for the 8-h colchicine treatment (15.62%).     

Destruction and reorganization of the receptor membrane in labellar chemosensory cells of the blowfly. Recovery of responses to sugar after destruction

Recovery from destruction by sodium deoxycholate (DOC) was studied with the receptor membrane of the blowfly, Phormia regina. The recovery can be divided into two processes, colchicine dependent and colchicine independent. The colchicine-dependent process was completely depressed by pretreatment with colchicine at 5 mM for 2 min (partially at 0.1 mM for 10 min), but the colchicine-independent one persisted. Vinblastine also caused depression but lumicolchicine did not. Records of responses obtained from the DOC-treated sugar receptor showed long response latencies that gradually became indistinct with recovery. Colchicine also affected this change in response latency after the DOC treatment. These results suggest that the colchicine-dependent recovery process is related to microtubules in the distal process of the receptor cell. The recovery time course and the change in response latency could be quantitatively explained by the simple assumptions that DOC underwent desorption from the receptor membrane (colchicine-independent recovery process) and that regeneration of the disrupted distal process of the receptor cell accompanied recovery in the number of available receptor sites (colchicine-dependent recovery process).     

24-表油菜素内酯对低氧胁迫下黄瓜根系抗氧化系统及无氧呼吸酶活性的影响

用营养液水培,研究了根际低氧胁迫下24-表油菜素内酯(EBR)对2个抗低氧能力不同的黄瓜品种根系中抗氧化系统及无氧呼吸酶活性的影响。结果表明,在低氧胁迫下,EBR处理显著提高了低氧胁迫下2品种黄瓜幼苗根系SOD、POD及ADH活性,降低了O2-·、H2O2和MDA含量、LDH活性及‘中农八号’根系PDC活性,而对‘绿霸春四号’根系PDC及2个品种CAT活性无明显影响,表明外源EBR处理通过促进低氧胁迫下根系中抗氧化酶和ADH活性的提高,降低LDH活性及ROS含量,增强植株抗低氧胁迫的能力。     

Immunofluorescence demonstration of tubulin and actin in estrogen-induced rat prolactinoma cells in vitro : Alteration of their distribution after bromocriptine, colchicine and cytochalasin B treatments

Cultured cells in vitro from estrogen-induced rat prolactin-secreting adenomas (prolactinomas) were examined by indirect immunofluorescence microscopy for the distribution of cytoskeletal proteins and alterations of cytoskeleton after treatment with bromocriptine, colchicine and cytochalasin B (CB). After 8 days in culture, prolactinoma cells were well expanded and developed cytoplasmic processes were seen. The cytoplasmic microtubules were observed as fine reticular networks radiating from perinuclear portions toward the cell periphery when decorated with an antibody against tubulin. On the other hand, the actin filaments showed diffuse and spotty distribution when detected with an anti-actin antibody. Contaminated fibroblasts showed a reticular distribution of microtubules and a parallel array of actin cables which corresponds to "stress fibers" throughout the cytoplasm. After treatment with bromocriptine, the reticular distribution of microtubules in prolactinoma cells changed into a coarse and sparse pattern, which was identical with the changes in the distribution of tubulin after treatment with colchicine. On the other hand, distribution of actin was not affected by bromocriptine. Bromocriptine treatment did not alter the distribution of microtubules and actin filaments in fibroblasts, whereas colchicine changed the distribution of microtubules in both prolactinoma cells and fibroblasts. CB treatment changed the localization of actin filaments in both kinds of cells. These in vitro studies indicated bromocriptine would selectively affect the cytoplasmic microtubular system of prolactinoma cells.     

Antioxidant properties of colchicine in acute carbon tetrachloride induced rat liver injury and its role in the resolution of established cirrhosis

Antioxidant and antifibrotic properties of colchicine were investigated in the carbon tetrachloride (CCl(4)) rat model. (1) The protective effect of colchicine pretreatment on CCl(4) induced oxidant stress was examined in rats subsequently receiving a single lethal dose of CCl(4). Urinary 8-isoprostane, kidney and liver malondialdehyde and kidney glutathione levels increased following CCl(4) treatment, but only the rise in kidney malondialdehyde was significantly inhibited by colchicine pretreatment. Serum total antioxidant levels were significantly higher in the colchicine pretreatment group. (2) The long term effects of colchicine treatment on CCl(4) induced liver damage were investigated using liver histology and biochemical markers (hydroxyproline and type III procollagen peptide). Co-administration of colchicine with sub-lethal doses of CCl(4) over 10 weeks did not prevent progression to cirrhosis. However, rats made cirrhotic with repeated CCl(4) challenge and subsequently treated with colchicine for 12 months, all showed histological regression of cirrhosis. (3) The antioxidant effect of colchicine in vitro was evident only at very high concentrations compared to other plasma antioxidants. In summary, colchicine has only weak antioxidant properties, but does afford some protection against oxidative stress; more importantly, long term treatment with this drug may be of value in producing regression of established cirrhosis.     

Electron microscope investigation of the evolutionary stages of the trophozoite of Didymophyes gigantea (Sporozoa, Gregarinida). III. The fine structure of the epicyte with emphasis on the contractile elements (author's transl)

The fine structure of the epicyte of D. gigantea was investigated. The motility of the gregarine and the contractile elements are described. Four essential types of movements can be observed in this gregarine: (1) rolling up and pendular movements, (2) locomotion by gliding forward, (3) cytoplasmic streaming (Fig. 1), (4) peristaltic contractions (Fig. 2) which seem to be accompanied by the contraction of annular myonemes (Fig. 2). The epicyte is formed by the folding of the parasitic cell wall which is made from three membranes (Figs. 3 and 4). At the top of each fold one can see apical struts between the outer and middle membrane and apical filaments under the inner membrane (Fig. 3). In addition, the epicytic folds are covered by a cell coat which is made from tubular structures (Fig. 5). At the base of the epicytic folds can be observed the basal lamina (Fig. 3) composed of very fine fibrillar material with an average thickness of 2.5 nm (Fig. 6). These fibrils are oriented in the longitudinal axis of the gregarine. Beneath the epicytic fold in the ectoplasm are found the annular myonemes with a width of up to 0.5 micrometers (Fig. 7). They are composed of many fine fibrils with an average thickness of 5 nm. In young trophozoites, the myonemes also contain microtubuli (Fig. 8). Between the epicytic folds, the cell wall is interrupted by three different types of vesicles: the vesicles with an electrondense content (Fig. 9), the three-membranous vesicles (Fig. 10), and the hose-shaped vesicles (Fig. 11). Glycerol-extraction of the parasites was performed in order to define the contractile structures. After extraction the annular myonemes are difficult to recognize (Fig. 13). When ATP is added, the gregarine does not contract but the myonemes reappear after 3 to 4 min (Fig. 14). Differences can also be observed in the myoneme structure using electron microscopy: After extraction, the myonemes are composed of a very limp fibrillar network (Fig. 15) which becomes very dense after the action of ATP (Fig. 16). Glycerol extraction does not disturb either the apical struts and apical filaments or the fibrils of the basal lamina (Figs. 15--17). In addition, cytoplasmic fibrillar structures appear after glycerol extraction (Figs. 15 and 16). The experimental and electron microscope results indicate that the motility of the gregarine depends upon four different systems: (1) the ectoplasmic annular myonemes, (2) the apical structures in the undulating epicytic folds, (3) the cytoplasmic fibrils, and (4) the basal lamina.     

Improved production of doubled haploids of winter and spring triticale hybrids via combination of colchicine treatments on anthers and regenerated plants

Double haploids (DH), obtained during androgenesis in vitro or by genome diploidisation in regenerated haploids, are one type of basic materials used in triticale breeding programmes. The aim of this study was to improve DH production by a combination of colchicine treatment methods on a sample of five winter and five spring triticale hybrids. Colchicine was applied in vitro either in the C17 medium to induce embryo-like structures (ELS) or in the 190-2 medium for green plant (GP) development. Regenerants which remained haploid were immersed in a colchicine solution either when placed on the medium prior to transferring to soil or when growing in pots, followed by the application or absence of cooling. Colchicine treatment during anther culture affected neither ELS nor GP development, but significantly increased the number of DH plants in comparison to spontaneous chromosome doubling. The highest efficiency was recorded when colchicine was applied in the induction medium (55%) versus the regeneration medium (44.5%) or no colchicine treatment (30%). The effectiveness of chromosome duplication in haploid plants ranged from 32 to 64.5% and it was the highest for the treatment on the medium followed by cooling. Individual hybrids differed regarding their capability of regeneration and chromosome doubling, which were consistent only to a low or moderate extent. However, taken together, winter and spring hybrids did not differ significantly. Combined colchicine application resulted in a high yield of DH production, 82.6% for all triticale hybrids, and can provide a considerable number of fertile DH lines for triticale breeding programmes.     

Microtubules, colchicine, and lymphocyte blastogenesis.

We have studied the time course of disassembly of microtubules of resting and stimulated mouse lymphocytes caused by the drug colchicine, as well as the effect of this compound on DNA and RNA synthesis of human and mouse lymphocytes. Fine-structure studies with the electron microscope showed a great increase in number of microtubules resulting from stimulation of mouse lymphocytes by the mitogenic lectin Con A. The presence of a network of microtubules was demonstrated in resting lymphocytes by use of the technique of immunofluorescence; this technique was not effective for the study of the microtubules of stimulated lymphocytes in the blast stage. The disappearance of microtubular networks in some cells (approximately 25%) was caused by the protocol of colchicine treatment used in many laboratories (30 min at 10(6) M); a 6- to 8-h treatment was required to cause all cells to lose their microtubules. It is indicated in these findings that there is need for extreme caution in implicating microtubule disruption as the cause of certain colchicine effects, such as that on the Con A-induced inhibition of receptor-ligand migration. The addition of colchicine to stimulated cells at varying times of culture caused marked inhibition of DNA synthesis provided that sufficient time (approximately 20 h for maximum inhibition) elapsed between addition of the drug to the stimulated culture and assay of DNA synthesis. Our data on the time course of inhibition of DNA synthesis by alpha-methyl mannoside (alpha MM) and by colchicine do not exclude the possibility that the latter compound may act partially by affecting the commitment of stimulated lymphocytes to DNA synthesis but they show that it can inhibit well after commitment is complete. The later the time of assay of thymidine incorporation, the more disparate were the curves relating the effects of alpha MM and colchicine to DNA synthesis of human cells. In the case of mouse splenic lymphocytes, there was no resemblance between the time course of the alpha MM and of the colchicine effects. Synthesis of RNA after 12 h of culture of stimulated human lymphocytes was also sensitive to colchicine.     

阿拉善荒漠连续放牧对三趾跳鼠巢区的影响

三趾跳鼠(Dipus sagitta)是内蒙古西部荒漠草原和沙地的优势害鼠,巢区研究对了解探讨其繁殖策略和种群动态规律有着重要的理论和实践意义。本研究通过2012~2014年的野外标志重捕实验,对三趾跳鼠的巢区在连续放牧和禁牧条件下的变动特征进行了分析。结果表明,连续放牧区雌性三趾跳鼠在6月的巢区范围显著大于禁牧区(P0.05);禁牧区雄性三趾跳鼠的巢区仅在7月份显著大于连续放牧区(P0.05),其他季节没有显著差异(P0.05)。放牧对雄性巢区无显著影响(P0.05),但连续放牧使雌性三趾跳鼠巢区范围显著大于禁牧区。雌性与雄性三趾跳鼠巢区均与一年生草本生物量存在正相关关系。这些结果表明,相对于雄性三趾跳鼠来说,雌性对由放牧导致的食物资源变化比较敏感,这种现象在哺乳期更为显著;繁殖是影响雄性三趾跳鼠巢区的因素之一。因此,保卫资源假说的理论可能是放牧导致三趾跳鼠巢区变化的重要原因之一。     

‘Switching-off’ of an auditory interneuron during stridulation in the acridid grasshopperChorthippus biguttulus L.

Summary In freely moving grasshoppers of the speciesChorthippus biguttulus compound potentials were recorded from the neck connectives with chronically implanted hook electrodes. The spikes of one large auditory interneuron, known as the G-neuron (Kalmring 1975a, b) were clearly distinguishable in the recordings and the neuron was identified by its physiology and morphology. In quiescent grasshoppers the G-neuron responds to auditory and vibratory stimuli, but responses to both stimuli are suppressed during stridulation in males (Fig. 1, top, Fig. 7). When a male's wings were removed so that the stridulatory movements of its hindlegs produced no sound, the suppression of the G-neuron response still occurred (Fig. 1, bottom). When proprioceptive feedback from the hindlegs was reduced, by forced autotomy of the legs, the switching-off remained incomplete (Fig. 3) (production of stridulatory patterns was inferred on the basis of electromyograms from the relevant thoracic musculature). Imposed movement of the hindlegs, on the other hand, suppressed the G-neuron response in a graded fashion, depending on the frequency of the movement (Figs. 4 and 5). These experiments suggest that the switching-off is brought about by a combination of proprioceptive feedback and central efferences. The switching-off phenomenon could either protect the grasshopper's auditory pathway from undesired effects of overloading by its own intense song (e.g. self-induced habituation as described by Krasne and Wine 1977) and should therefore apply for most auditory neurons. Alternatively it could prevent escape reflexes from being triggered by stridulatory self-stimulation and consequently might apply only for neurons involved in such networks (as the G-neuron might be).     

镉诱导的茶树苗膜脂过氧化和细胞程序性死亡

在含镉的营养液中,茶树幼苗生长受到抑制。随培养时间延长,膜脂过氧化产物丙二醛含量持续升高;超氧物岐化酶（SOD）和过氧化氢酶（CAT）活性初期升高,而后分别在第4天和第2天开始下降。镉胁迫的第5-7天,一部分细胞陆续发生程序性死亡。其特征是：线粒体聚集于核周围,个数增加,嵴发达,而后衰亡。核仁消失,染色质凝结在核膜边缘,核萎缩,外层核膜局部扩张,形成胀泡。核以外溢、出芽和崩裂三种方式溃解。核是最后消亡的细胞器。程序性死亡的细胞局限于某些区域。镉胁迫下,幼苗膜脂过氧化可能是诱发PCD的主要原因。     

大花蕙兰四倍体的离体诱导和鉴定

采用组织培养方法研究了秋水仙素处理对大花蕙兰类原球茎增殖、分化和四倍体诱导的效应.结果表明,秋水仙素处理明显抑制类原球茎的增殖和分化,提高类原球茎褐变率和死亡率;在试验浓度和时间范围内,秋水仙素浓度越高,处理时间越长,抑制作用越强,解除抑制作用所需的继代次数越多.所有秋水仙素处理均能诱导出四倍体,但不同处理的四倍体诱导率不同,当处理浓度为0.05%、时间为5 d时,四倍体诱导率最高,为23.7%.二倍体及其四倍体的气孔保卫细胞长度和气孔密度均存在极显著的差异.四倍体植株比二倍体矮,茎部较粗壮,株型紧凑,叶片颜色浓绿、质地变硬.秋水仙素处理和组织培养相结合是创建大花蕙兰多倍体的有效方法.     

Podophyllotoxin as a probe for the colchicine binding site of tubulin.

The binding of [3H]podophyllotoxin to tubulin, measured by a DEAE-cellulose filter paper method, occurs with an affinity constant of 1.8 X 10(6) M-1 (37 degrees at pH 6.7). Like colchicine, approximately 0.8 mol of podophyllotixin are bound per mol of tubulin dimer, and the reaction is entropy-driven (43 cal deg-1 mol-1). At 37 degrees the association rate constant for podophyllotoxin binding is 3.8 X 10(6) M-1 h-1, approximtaely 10 times higher than for colchicine; this is reflected in the activation energies for binding which are 14.7 kcal/mol for podophyllotoxin and 20.3 kcal/mol for colchicine. The dissociation rate constant for the tubulin-podophyllotoxin complex is 1.9 h-1, and the affinity constant calculated from the ratio of the rates is close to that obtained by equilibrium measurements. Podophyllotxin and colchicine are mutually competitive inhibitors. This can be ascribed to the fact that both compounds have a trimethoxyphenyl ring and analogues of either compound with bulky substituents in their trimethoxyphenyl moiety are unable to inhibit the the binding of either of the two ligands. Tropolone, which inhibits colchicine binding competitively, has no effect on the podophyllotoxin/tubulin reaction. Conversely, podophyllotoxin does not influence tropolone binding. Moreover, the tropolone binding site of tubulin does not show the temperature and pH lability of the colchicine and podophyllotoxin domains, hence this lability can be ascribed to the trimethoxyphenyl binding region of tubulin. Since podophyllotoxin analogues with a modified B ring do not bind, it is concluded that both podophyllotoxin and colchicine each have at least two points of attachment to tubulin and that they share one of them, the binding region of the trimethoxyphenyl moiety.     

Discoupling of excitation-contraction processes by urea treatment of frog skeletal muscle

On exposure (E) of frog semitendinosus muscle to 400 mmol/l urea (U) in sodium chloride Ringer's solution, the tension development to isoK+ solutions decreased, while in choline chloride Ringer it increased. On quick removal (R) of urea, always a block of excitation-contraction (E-C) coupling occurred accompanied by transient or persistent swelling of fibres and a similar but definite decrease of their resting membrane potential (Fig. 2). Muscle contraction could be elicited by caffeine even after UER-treatment but then only the slow tension increase (second phase of normal caffeine contraction) occurred (Fig. 3a). The fast tension increase to caffeine (first phase) could be restored if after UER-treatment 5 mmol/l mannitol (Fig. 3b), a 20 min treatment with choline chloride (Fig. 4a) or sodium isethionate (Fig. 4b) Ringer's solution of double osmolarity were applied. Caffeine contraction could not be elicited when sodium chloride Ringer's solution of double osmolarity was used under similar conditions (Fig. 5). E-C block to isoK+ solution persisted in all these experiments. E-C coupling could partially be restored by short treatment of muscle with caffeine (Figs 6a, b).     

外源H2O2对湖北海棠根系线粒体膜透性和细胞核DNA的影响

用0．024％（V／V）H2O2处理湖北海棠（Malus hupehensis Rehd．）实生苗根系,30min后,根系线粒体膜通透,陛明显增大,线粒体膜电位（△ψm）和线粒体内Cyt c／a吸光度比值下降;60min后,根系细胞核DNA发生片段化降解。经荧光染料吖啶橙染色后,可见H2O2处理60min以上的根系压片中出现了清晰致密的黄绿色荧光宽,呈现出细胞程序,陛死亡（PCD）的特征。     

干旱过程中露花叶片PEPCase酶蛋白量的变化

干旱和盐胁迫使露花叶片中PEPCase活性水平提高数倍。经双相电泳和蛋白免疫印溃(Western immunoblot)发现干旱后露花叶片中PEPCase蛋白量有明显增多。火箭免疫电泳定量分析结果表明,干旱过程中,PEPCase活性水平的提高主要基于其酶蛋白量的增加。干旱引起的这些变化是可逆的,在恢复供水一定时间后可以完全恢复。RubisCO蛋白量在干旱处理的露花叶片中明显减少,恢复供水后又回复到原初水平。PEPCase蛋白量增多,RubisCO蛋白量减少可能是露花叶片CAM途径诱导的生化特征之一。露花叶片中PEPCase活性水平及蛋白量受到叶片发育状况的控制,干旱引起的这两者的变化在中部成熟叶片中最明显,在幼嫩叶片中最小。