Studies on Haematococcus pluvialis for improved production of astaxanthin by mutagenesis

Cultures of Haematococcus pluvialis were exposed to mutagens like u.v. and EMS (ethyl methanesulphonate). The results showed that the survival rate decreased with the increase in u.v. exposure time and increase in EMS concentration. These mutants were further screened using inhibitors of the carotenoid biosynthetic pathway viz. diphenylamine (15–90 M), nicotine (160–320 M) and compactin (1.5–3.0 M). The mutants thus obtained showed early enhanced (2.2–3.2-fold) astaxanthin accumulation and also exhibited higher lycopene cyclase activity.     

Saccharomyces cerevisiae 2-mum DNA. An analysis of the monomer and its multimers by electron microscopy.

Summary The non-tandem inverted duplication in the 2-m DNA of Saccharomyces cerevisiae has a length of 0.19 m and is located asymmetrically along the molecule. The majority of the dumb-bell structures that are formed upon denaturation and selfannealing of the 2-m monomer consists of the renatured inverted duplication sequences as double stranded stem and two single stranded loops of 0.67 m±0.06 m (S-loop) and 0.86 m±0.05 m (L-loop) length. Two additional size classes which comprised 5–10% of the measured molecules had contour lengths of around 1.7 m and 2.1 m. The smaller dumb-bells contained two S-loops and the larger dumb-bells contained two L-loops as was shown by heteroduplex mapping with an HindIII fragment from the L-loop. Two models which assume illegitimate or site specific recombination, are presented to explain the generation of double S-loop and double L-loop molecules. At least part of the 4-m and 6- circular molecules present in the yeast supercoiled DNA fraction are shown to be dimers and trimers of 2-m monomers, but often with inverted loop segments most probably due to intramolecular recombination between sequences of the inverted duplication.2-m DNA is used to indicate the supercoiled DNA fraction although in our measurements the average monomeric length is 1.9 mPart of this work has been presented at the Conference: The Genetics and Biogenesis of Chloroplasts and Mitochondria, Munich, August, 1976     

Effects of a new class I antiarrhythmic drug bidisomide on canine ventricular arrhythmia models

The antiarrhythmic and direct cardiovascular effects of a new antiarrhythmic agent, bidisomide, -{2-[acetyl(1-methylethyl)amino]ethyl}--(2-chlorophenyl)-1-piperidinebutanamide, were investigated. To determine the antiarrhythmic effects, spontaneously occurring adrenaline-, digitalis-and two-stage coronary ligation-induced arrhythmias were used. Bidisomide suppressed these three arrhythmia models. The antiarrhythmic plasma concentration, IC₅₀, of bidisomide for digitalis-induced arrhythmia was 22.1 g/ml, and those calculated for intravenous bidisomide in 24 h and 48 h coronary ligation-arrhythmias were 15.1 and 11.6 g/ml and that calculated for oral bidisomide in 24 h coronary ligation-arrhythmia was 5.4 g/ml and that for adrenaline induced arrhythmia was 58.7 g/ml. In the blood perfused sinoatrial node and papillary muscle preparations, bidisomide decreased the sinoatrial rate and contractile force and increased the intraventricular conduction time and coronary blood flow. These results indicate that bidisomide is similar to other class I antiarrhythmic agents such as pirmenol and KW-3401 in its antiarrhythmic profile and is expected to become a clinically useful antiarrhythmic drug.     

Zur Feinstruktur der Elektrozeptorepidermis der Mormyridae (Teleostei,Pisces)

Zusammenfassung Die schwachelektrischen Mormyridae haben eine dreischichtige Epidermis, deren innere Schicht aus nur etwa 0,22 m dicken sechseckigen Zellen von ca. 60 m Durchmesser besteht. Die etwa 2 m dicken, linsenförmigen Kerne von 7,6 m Durchmesser liegen am Zellrand. Die Zellen sind zu Säulen aufgeschichtet. Ihr Rand ist ausgezackt und dort, wo er die Säulengrenze erreicht, auf etwa 0,34 m verdickt. In der Nähe der Säulengrenzen sind die Zellen über Desmosomen mit den Nachbarn in der eigenen und in der angrenzenden Säule verbunden. Diese Epidermisschicht ist auf die Körperpartien beschränkt, in denen auch Elektrorezeptoren ausgebildet sind.Die beiden anderen Epidermisschichten haben den üblichen Aufbau einer Fischepidermis, abgesehen vom Fehlen der Becherzellen.

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Ultrastructure of the electroceptor epidermis of the Mormyridae (Teleostei, Pisces)

Summary The weakly electric fish of the family Mormyridae have a three layered epidermis, with a medium layer consisting of hexagonal cells of only 0.22 m in thickness and about 60 m in diameter. The lens-shaped nuclei are about 2 m thick and 7.6 m in diameter and are situated near the border of the cells. The cells are piled up to hexagonal columns. Their margin is serrate and where it reaches the boundary of the column, it has a thickness of about 0.34 m. Close to the boundaries of the columns, the cells are linked to their neighbours within the column and of the adjoining column by desmosomes. This layer of the epidermis is confined to those regions of the body surface which also contain electroreceptors.The other layers of the epidermis have a structure as usual in fish, except for the lack of goblet cells.

     

Plant regeneration from embryo-derived callus of oil palm – the effect of exogenous polyamines

Regeneration in oil palm was achieved through somatic embryogenesis/organogenesis from embryo-derived callus. Callus was induced from mature embryos of the cross 281 (D)×18 (P) on modified MS medium supplemented with 2,4-D (113.12 M) and 2-iP (14.76 M). The embryogenic calluses obtained were transferred to Blaydes medium supplemented with 2,4-D (0.045 M) and one of the following growth regulators: TDZ (4.54 M), zeatin riboside (2.85 M), putrescine (1 mM) and spermine (100 M). Secondary somatic embryogenesis was found to occur in media supplemented with polyamines. The efficiency of formation of somatic embryos, secondary somatic embryos and shoot meristemoids were significantly higher in putrescine containing medium. Histological studies were also undertaken.     

Regeneration of plantlets through organogenesis in the Himalayan yellow poppy,Meconopsis paniculata

Plantlet formation through organogenesis in callus cultures of Himalayan yellow poppy,Meconopsis paniculata D.Don (Prain), a threatened taxon of ornamental value, is described. Hypocotyl segments from 3-month-old laboratory-raised seedlings produced callus on agar-solidified Murashige and Skoog medium (MS) containing 10 M -naphthaleneacetic acid and 1 M kinetin. Shoots differentiated best from callus on MS containing 10 M indolebutyric acid (IBA) and 1 M 6-benzyladenine. The regenerated shoots rooted best on MS medium containing 10 M IBA. From seed germination to differentiation of plantlets through the two-step organogenesis process required 28–29 weeks.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - FAA formalin-acetic acid-alcohol - BA 6-benzyladenine - IAA indole-acetic acid - IBA indolebutyric acid - GA₃ gibberellic acid - NAA -naphthaleneacetic acid - RH relative humidity     

Effects of mefluidide and dicamba on in vitro growth and embryogenesis ofDactylis glomerata (orchard grass)

The effects of N-(2,4-dimethyl-5-(((trifluoromethyl)sulfonyl)amino)phenyl)acetamide (mefluidide) and 3,6-dichloro-o-anisic acid (dicamba) on in vitro growth and somatic embryogenesis ofDactylis glomerata L. (orchard grass) were studied using suspension cultures and explanted leaf bases. All experiments employed modified Schenk and Hildebrandt medium amended with concentrations of dicamba ranging from 15 to 120 M (SH-15 to SH-120) and of mefluidide ranging from 1 to 100 M. SH medium without either growth regulator was used for embryo germination. Embyro production in suspension cultures with SH-30 medium plus 3 g/L casein hydrolysate was significantly reduced by 1 M mefluidide. Only 15% of these embryos germinated and produced plants compared to 84% from controls. Growth, as measured by dry weight, was significantly reduced by 50 or 100 M mefluidide. The number of embryos formed on leaf sections was significantly reduced by 20 or 25 M mefluidide. Embryos that formed with 10 M or more mefluidide were callused on both SH-15 and SH-30 media. Shoot formation was inhibited from individual embryos and embryo/callus masses that developed on either SH-15 or SH-30 medium containing 5 M or more mefluidide. Radicle emergence was significantly reduced with 10 M mefluidide regardless of 15 or 30 M dicamba. Histological examination revealed that mefluidide inhibited both shoot and root meristem development with shoot development being the more sensitive. Inhibition of both was independent of dicamba concentrations. Shoot formation was also reduced from embryos that had developed on SH-30 medium without mefluidide when transferred to medium containing mefluidide without dicamba.     

Influence of sarmesin on some dopamine-related types of behaviour

Summary The present study was undertaken to investigate the effects of sarmesin, an analogue of [Sar¹] angiotensin II (ANG II) where the tyrosine hydroxyl group in position 4 is methylated, on dopamine (DA)-related paradigms: locomotor and exploratory behaviour as well as apomorphine (3 mg/kg, ip)-induced stereotypy in rats. Sarmesin (0.5 and 1 g, icv) significantly decreased ambulation and rearing movements, and blocked the inhibitory effect of ANG II (0.1 g) on both types of activity. Sarmesin induced biphasic effects on apomorphine-induced stereotypy depending on the dose increase (0.5 and 5 g, icv) and decrease (10 g). Moreover, sarmesin (5 g) blocked the inhibitory effect of ANG II (2 g, icv) on apomorphine stereotypy. Taken together, these results suggest that sarmesin might interact with AT₁ and AT₂ receptor subtypes. The results further confirm the statement for ANG II-DA interaction in brain structures involved in these types of behaviour.     

A comparison of BA,zeatin and thidiazuron for adventitious bud formation from Picea glauca embryos and epicotyl explants

Picea glauca (white spruce) zygotic embryos and one-week-old-seedling epicotyl explants were placed on either Woody Plant Medium (WPM) or half-strength Schenk & Hildebrandt (1/2S&H) medium supplemented with varying levels of benzyladenine (BA) (0.1, 1.0, 10, 50, 100 M), zeatin (10, 50, 100 M) or thidiazuron (TDZ) (0.01, 0.1 M). In addition to differences in the number of buds induced at three months on the two media, buds induced on WPM were visually more uniform, less vitrified and elongated faster. On 1/2S&H supplemented with BA, maximum bud induction from embryos occurred on 1.0 M BA with 0.01 M TDZ with higher BA concentrations inhibitory to bud induction. In contrast, on WPM there was little difference in the number of buds induced from embryos placed on 10, 50 and 100 M BA with or without TDZ. One-week-old-seedling epicotyl explants required higher BA levels on 1/2S&H, as bud induction at three months was greatest at 10 M BA. On WPM, as with the embryos, there were only minor differences in the number of buds induced from epicotyl explants on the various BA levels. Zeatin was more effective at inducing buds than BA with both media. From embryos, bud induction was greatest on 50 or 100 M zeatin without TDZ and 50 or 100 M zeatin with or without TDZ on 1/2S&H and WPM respectively. From epicotyl explants on 1/2S&H, there was little difference in the number of buds induced with the zeatin concentrations used, while with WPM, 50 and 100 M zeatin induced the greatest number of buds. Interestingly, with BA, the epicotyl explants needed a higher level than the embryos for maximal response, while with zeatin, the level was the same for both embryos and epicotyl explants. Long-term (six month) survival was higher on WPM than with 1/2S&H. Additionally, embryos had a higher percentage of genotypes surviving at six-months when compared with epicotyl explants. For overall survival and development of the buds, 50 M zeatin with 0.01 M TDZ was the best treatment tested.Abbreviations BA benzyladenine, 1/2S&H-half-strength Schenk & Hildebrandt medium - TDZ thidiazuron - WPM woody plant medium     

Lead‐related effects on rat fibroblasts

Lead (Pb) is an environmental toxicant that can induce structural and functional abnormalities of multiple organ systems, including the central nervous and the immune systems. The aim of this study was to evaluate the effects of extracellular Pb supplementation on the cellular content of the metal and on the proliferation and the survival of normal rat fibroblasts.We found that the concentration of Pb in the culture medium was 0.060 M and the normal Pb concentration in rat fibroblasts was 3.1 ± 0.1 ng/10⁷ cells. Then we exposed the cells to increasing concentration of Pb (as Pb acetate) from 0.078–320 M. We observed a dosedependent inhibition of cell proliferation after 48 h, which was already apparent at a concentration of 0.312 M (p = 0.122) and became statistically significant for concentration higher than 0.625 M (p = 0.0003 at 5 M). Cell proliferation was completely compromised at 320 M Pb total inhibition of cell proliferation.To investigate the mechanisms of Pbmediated inhibition of cell proliferation, we evaluated the occurrence of apoptosis in the same cells and found that cytosolic DNA fragments, hallmark of apoptotic cell death, increased significantly at Pb concentrations from 2.5–10.0 M. The occurrence of apoptosis was also confirmed by FACS analysis which showed the appearance of a subdiploid peak at Pb concentrations from 5–20 M. The distribution of cells in the cell cycle showed a dosedependent accumulation of cells in the G₀/G₁ phase mainly compensated by a decrease in the percentage of cells in the S phase. In conclusion, our results demonstrate that induction of apoptosis contributes to the Pbinduced inhibition of cell proliferation in rat fibroblasts.     

Measurements of manganese in thylakoids of Sinapis alba grown under high-light and low-light conditions

The manganese content of thylakoids and tissues was measured in leaves grown under high- and low-light conditions. Especially when grown in a nutrient medium enriched in manganese (20 M), the thylakoids contained large amounts of manganese, which could be removed by EDTA washing without impairment of the Hill reaction. The unremovable content of manganese was almost the same in thylakoids from plants grown in nutrient media of normal (2 M) and reduced (0.2 M) manganese content. Up to this limit of manganese content, Hill activity did not seem to be impaired. 1.2 atoms Mn per 100 molecules chlorophyll were found in low-light thylakoids and 1.6 atoms Mn in high-light thylakoids. This is similar to the behaviour of other electron transport components, the number of which is also decreased under low-light conditions. However, the decrease in the manganese content is not as striking as the decrease in, for example, the cytochrome f and ferredoxin content. This may be attributed to an invariable pool of manganese which is not involved in the oxygen evolving system. Alternatively, if all of our measured manganese is involved in electron transport to PS II, this could indicate that in low-light chloroplasts the ratio of PS II/PS I components may be somewhat increased.

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Zusammenfassung Der Mangangehalt von Thylakoiden und Gewebe aus Starklicht- und Schwachlichtblättern wurde untersucht. Besonders bei Pflanzen, welche unter erhöhtem Manganangebot (20 M) angezogen wurden, besaßen die Thylakoide sehr viel Mangan, welches durch Waschen mit EDTA entfernt werden konnte, ohne die Hill-Aktivität zu beeinträchtigen. In Thylakoiden aus Pflanzen, welche unter normalem (2 M) und reduziertem (0,2 M) Manganangebot gewachsen waren, unterschied sich der nicht entfernbare Mangangehalt nicht sehr. Dies scheint die untere Grenze des Mangangehalts zu sein, bis zu welchem die Hill-Aktivität noch nicht beeinträchtigt wird. Schwachlicht-Thylakoide besitzen 1,2 Atome Mn pro 100 Chlorophyllmoleküle, während Starklicht-Thylakoide 1,6 Atome Mn pro 100 Chlorophyllmoleküle enthalten. Dies gleicht dem Verhalten anderer Komponenten des Elektronentransports, welche ebenfalls im Starklicht vermehrt vorkommen. Die Unterschiede im Mangangehalt sind jedoch geringer als die Unterschiede im Gehalt von z.B. Cytochrom f und Ferredoxin. Dies könnte auf einen konstanten Anteil von Mangan zurückzuführen sein, welcher nicht am wasserspaltenden System beteiligt ist. Wenn jedoch das gesamte gemessene Mangan am Elektronentransport zum PS II beteiligt ist, könnte dies ein Hinweis sein, daß in Schwachlicht-Chloroplasten sich das Verhältnis der PS II-/PS I-Komponenten etwas vergrößert.

     

Rapid plant regeneration from callus cultures of Plumbago zeylanica

Rapid plant regeneration was achieved in callus cultures derived from leaf and stem explants of Plumbago zeylanica Linn. on MS basal medium supplemented with 4.44 M 6-BA, 1.42 M IAA and 3% (w/v) sucrose. The rate of shoot bud regeneration was positively correlated with the concentration of growth regulators in the nutrient media. The leaf explants were more responsive (82.3%) than the stem explants on medium containing 1.42M IAA in combination with 4.44 M BA. The rate of regeneration was found to maintain the same level for 12 months without loss of vigour. Rooting of the differentiated shoots was achieved in media having 0.57 M IAA with 2% (w/v) sucrose within 10 days of culture. Regenerated plantlets were transferred to soil which grew normally with a survival rate of 90%. This protocol may help in the conservation of the species and selection of variants that may be induced to widen the genetic base of the genus.     

Three dimensional reconstruction of the X-Y pair during pachytene in the rat (Rattus norvegicus)

The ultrastructure of the X-Y pair from rat spermatocytes has been reconstructed sterically by the study of serial sections. The X-Y pair of the rat at early pachytene contains two dense cores, a long and a short one, which form a synaptonemal complex 1.7 long at their common end. The long core (10.6 ) and the short core (4.5 ) correspond to X and Y, respectively. There is no RNA histochemically detectable in the X-Y pair. Nucleoli are independent of the X-Y pair. A low number of cells show nucleoli very near the X-Y pair but no continuity exists between these structures.     

Asymmetric diffusion into the postsynaptic neuron: An extremely efficient mechanism for removing excess GABA from synaptic clefts on the Deiters' neurone plasma membrane

Microdissected Deiters' neuron plasma membranes have been used for studying the passage of GABA through the membrane both in the inward and outward direction. Working with 0.2 mM GABA in the compartment simulating the outside of the neurone and with 2.0 mM GABA in the one simulating the inside we found a net transport of GABA towards the inside. This mechanism does not require a Na⁺ ion gradient across the membrane. The nature of the transport process involved was studied by determining the rate of [³H]-GABA inward passage as a function of GABA concentration (1 nM–800 M) on the outward side of the membrane. The results have shown that until 50 M a diffusion process (v=D₁×C, where D₁=3.1×10^–11 1/m²×sec) is the sole mechanism involved. Above 50 M a second diffusion process is activated v=D₂×(C–50×10^–6), where D₂=2.8×10^–11 1/m²×sec. Taking in account both inward and outward directed diffusion, one can calculate 16 M as the equilibrium concentration of GABA on the outward side of the membrane. From a kinetic point of view, these diffusion processes are able to reduce GABA concentration in a synaptic cleft from 3 mM to 20 M within 3 sec. These diffusion systems are discussed as extremely efficient in removing the excess of released GABA in the synaptic cleft.     

Induction of apoptosis by gemcitabine in BG-1 human ovarian cancer cells compared with induction by staurosporine, paclitaxel and cisplatin

A variety of chemotherapeutic agents induce cell death via apoptosis. We had shown previously that gemcitabine (2,2-difluorodeoxycytidine) induced an atypical apoptosis in BG-1 human ovarian cancer cells; therefore, further studies were conducted to characterize more precisely gemcitabine-induced apoptosis in BG-1 cells compared to a general inducer of apoptosis, staurosporine. BG-1 cells exposed to 0.5, 1.0 and 10 M gemcitabine for 8 h, or staurosporine (1.0 M) for 6 h, exhibited high molecular weight DNA fragmentation (50 kbp); however, only staurosporine treatment produced internucleosomal DNA fragments (200 bp) in a laddered pattern on the agarose gel. Staurosporine (1.0 M) rapidly induced phosphatidylserine plasma membrane translocation that increased linearly with time as measured by annexin V-FITC binding, and similar kinetics were observed for caspase activation by staurosporine in BG-1 cells. In contrast, 10 M gemcitabine increased phosphatidylserine expression in a small fraction of cells (5–10%) vs. untreated controls over the course of 48 h and significant caspase activity was detected within 12 h of drug exposure. Time-lapse video microscopy of BG-1 cells exposed to 1.0 M staurosporine or 10 M gemcitabine for up to 72 h showed that the morphologic changes and kinetics of cell death induced by these agents differed significantly. We also evaluated the apoptosis induced by paclitaxel (a mitotic poison) and cisplatin (an agent not dependent on cell cycle functions) in BG-1 cells by these methods because these drugs are used clinically to treat ovarian cancer. Our findings demonstrate that the kinetics of apoptotic cell death induced by gemcitabine and other chemotherapeutic agents should be taken into account when designing treatment strategies for ovarian cancer.     

Zinc protects against arsenic-induced apoptosis in a neuronal cell line,measured by DEVD-caspase activity

Acute and chronic arsenic exposure results in toxicity in humans and causes many neurological and other manifestations. For the first time the present study reports that zinc decreases arsenic-induced apoptosis and also confirms a single report of apoptosis induced by arsenic in a neuronal cell line. Apoptosis measured by DEVD-caspase activity peaked between 10 M and 20 M of arsenic trioxide. Higher concentrations of arsenic up to 40 M caused increasing cell death with diminishing DEVD-caspase activity. The beneficial effect of zinc was proportional to its concentration with a significant decrease in arsenic-induced DEVD-caspase activity at 50 M and 75 M zinc (P<0.05). This finding may be of therapeutic benefit in people suffering from chronic exposure to arsenic from natural sources, a global problem especially relevant to millions of people on the Indian subcontinent.     

Adventitious bud formation and shoot development from in vitro leaves of Vitis x Muscadinia hybrids

High levels of regeneration were obtained from young leaves excised from axillary shoots in proliferating nodal cultures of several Vitis x Muscadinia hybrids. Best results were obtained when the explants were cultured on solidified half-strength Murashige and Skoog medium supplemented with 8.9 M 6-benzyladenine and 0.05 M 1-naphthaleneacetic acid. Though variation was observed among the hybrids, the procedure used does not seem to be genotype-specific as all the hybrids and cultivars tested could regenerate. Scanning electron microscopy observations and histological studies carried out during the development of adventitious shoot organogenesis revealed that the promeristem initiation occurred in the outer cell layers near the wounded area of the petiolar stub.Abbreviations BA 6-benzyladenine - NAA 1-naphthaleneacetic acid     

Effect of thidiazuron on vegetative tissue-derived somatic embryogenesis and flowering of bamboo Bambusa edulis

Current research on somatic embryogenesis of bamboo uses reproductive tissue as explants. However, it was hard to obtain the explant. Shoots of a local accession (3–4 m high) were used for multiple shoot production. In order to obtain embryogenic callus, nodal and internodal tissues from in vitro plantlets were placed on Murashige and Skoog (MS) medium supplemented with 9.2 M kinetin (KN), 13.6 M 2,4-dichlorophenoxyacetic acid (2,4-D), 0.1% (v/v) coconut milk, and 6% (w/v) sucrose. We studied the effects of sucrose and thidiazuron (TDZ) on callus proliferation. Optimal additives to the MS medium for embryogenic callus proliferation were 0.046 M TDZ, 13.6 M 2,4-D and 3% (w/v) sucrose. TDZ also promoted the germination of bamboo somatic embryos. The germination rate of the somatic embryos exceeded 80% on MS-based medium supplemented with 0.455M TDZ. Naphthaleneacetic acid (NAA) reduced germination. Well-developed plantlets were successfully transferred to soil. There was no albino mutant in subsequent culture. In vitro regenerants and potted plants flowered, but no seeds were produced.     

Clearance rates of bacteria-sized particles by freshwater ciliates,measured with monodisperse fluorescent latex beads

Summary Monodisperse fluorescent latex particles with diameters of 0.57 and 1.04 m have been used to measure the clearance rates of bacteria-sized particles by two ciliates from a Norwegian lake. The clearance rates by Epistylis rotans on these particles were in the range 0.23 to 1.26 l ind^-1h^-1, and by Strombidium sp. 0.26 to 0.90 l ind^-1 h^-1. The clearance rates varied with food level and temperature. Possibilities and limitations of the suggested method are discussed.