Zur Kompartimentierung der Synthese von Mono- und Sesqui-Terpenen des ätherischen Öls beiPoncirus trifoliata

Zusammenfassung In der Frucht vonPoncirus trifoliata liegen in der Außenschale Drüsenzellkomplexe, die ein monoterpenreiches ätherisches Öl mit geringem Anteil an Sesquiterpenen und O-haltigen Substanzen produzieren. Ähnlich aussehende Exkretzellkomplexe aus den Saftschläuchen enthalten hauptsächlich Sesquiterpenkohlenwasserstoffe (STKW) und O-haltige Komponenten und sehr wenig Monoterpenkohlenwasserstoffe (MTKW). Im Schalenöl konnten nach gaschromatographischer Trennung mit Hilfe der Massenspektrometrie 19 Komponenten identifiziert werden, im Saftschlauchöl 25.Elektronenmikroskopische Aufnahmen der jüngsten Drüsenzellen beider Drüsenkomplexe lassen erkennen, daß beide Terpenklassen wahrscheinlich hauptsächlich bzw. ausschließlich plastidär entstehen.Exogen angebotenes¹⁴CO₂ wird zunächst überwiegend in die MTKW eingebaut, erst später nimmt die Markierung der STKW und O-haltigen Komponenten stark zu. Über den Ferntransportweg angebotenes¹⁴C-Leucin führt anfangs zu einer starken Markierung der STKW und O-haltigen Komponenten, erst später verschiebt sich der Einbau etwas mehr in Richtung MTKW. Als Hauptursache für den differenten Einbau wird das Vorhandensein zweier Typen von Drüsenzellkomplexen mit unterschiedlichen Syntheseleistungen angesehen.Die aus dem¹⁴CO₂ in der Außenrinde gebildeten Assimilate werden zuerst in das MTKW-reiche Öl der Schalenexkretbehälter eingebaut. Die überwiegend STKW erzeugenden Saftschlauchbehälter werden erst später beliefert. Beim Leucinangebot über die Fruchtstiele scheint es gerade umgekehrt zu verlaufen. Die aufeinanderfolgenden Maxima der Ölproduktion in den beiden Drüsenzellkomplex-Typen und die Änderung des Komponentenspektrums ihres ätherischen Öls im Verlauf der Vegetationsperiode tragen ebenfalls zu einem je nach Jahreszeit unterschiedlichen Einbau in die MTKW und STKW bei.

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Compartmentation of mono- and sesqui-terpene biosynthesis of the essential oil inPoncirus trifoliata

Summary The fruit ofPoncirus trifoliata shows glandular cell complexes in the exocarp, which produce a volatile oil rich in monoterpenes but poor in sesquiterpenes and oxigenated compounds. The juice vesicles of the endocarp possess similar cell complexes mainly containing sesquiterpenes and oxigenated compounds, whereas monoterpenes only occur in small amounts. By the use of combined gas chromatography-mass spectrometry 19 components of the rind oil and 15 compounds of the endocarp oil could be identified.As demonstrated by electron microscopy the terpenes most probably are synthesized predominantly, if not exclusively in plastids. As shown by gasradiochromatography radioactive precursors (¹⁴CO₂ and¹⁴C-leucine) are incorporated into mono- and sesqui-terpenes to a different extent.This is due to two gland types producing essential oils of different composition with regard to their mono- and sesqui-terpene percentage. In fruit development the exocarp glands differentiate earlier than the endocarp glands do. The activity of exogenously applied¹⁴CO₂ first reaches the peripheral glands and later on appears in the interior glands. Depending upon the growth season, labelled leucine transported by the conducting tissues from lower plant parts leads to a high specific activity of the sesqui-terpenes and oxigenated compounds. It could be argued that in this instance the glands of the pulp are better provided with precursors than the exocarp glands. The successive maxima of essential oil production in both glandular complexes, and the changes in the concentration of individual oil constituents during the ontogeny of the fruit also contribute to different incorporation ratios of radioactive precursors into mono- and sesqui-terpenes.

     

Untersuchungen zur Physiologie der Carnivoren-Drüsen

Zusammenfassung Der Cl^--Transport durch isolierte Gewebescheiben aus dem die überdachten Drüsen tragenden Teil von Nepenthes-Kannen wurde untersucht. An der der Kannenaußenseite entsprechenden Fläche, deren cutinisierte Epidermis durch oberflächliches Anschneiden mit einer Rasierklinge entfernt wurde, nehmen die Gewebescheiben aus den Versuchslösungen Cl^- durch metabolischen Trägertransport auf, während die Cl^--Abgabe passiv ist. Die Cl^--Sekretion durch die drüsentragende Oberfläche hängt von der Bereitstellung energiereicher Phosphate durch den Stoffwechsel ab. Zwischen dem Chloridgehalt und der caseinspaltenden Aktivität des Kannensekretes konnte eine Korrelation nachgewiesen werden.

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Investigations on the physiology of the glands of carnivorous plantsIV. The kinetics of chloride secretion by the gland tissue of Nepenthes

Summary The transport of chloride in isolated tissue from Nepenthes pitchers was investigated using ³⁶Cl^-, an Aminco-Cotlove chloride-titrator for the determinations of Cl^- concentrations, and KCN and AsO ₄ ^- -as metabolic inhibitors.The tissue was brought in contact with different experimental solutions (=medium). The surface corresponding to the outside of the pitchers was cut with a razor blade to remove the cutinized epidermal layer. At this surface the Cl^- uptake from the medium is a metabolic process which depends on the Cl^--concentration of the medium in a manner that corresponds to the Michaelis-Menten kinetics. The Michaelis-constant of this transport step was 3×10^-2M. The Cl^--efflux into the medium, however, is a passive process.The opposite surface of the tissue slices (corresponding to the inside of the pitchers) carries the glands. The chloride secretion taking place here is also dependent on metabolism. In vitro it occurs even when a high gradient of chloride concentration has been set up between the medium and the solution which is in contact with the glands. In vivo the Cl^--concentration of the pitcher fluid and the amount of Cl^- per gram of tissue water are almost equal.The rôle of chloride in the physiology of Nepenthes is still under investigation, A correlation between the chloride content of the pitcher fluid and its enzymatic activity (Casein-test), however, could already be demonstrated.

     

Untersuchungen zur Physiologie der Carnivoren-Drüsen

Zusammenfassung Die Sekretion von ³⁶Cl^- durch Gewebescheiben, die aus dem drüsentragenden Teil von Nepenthes-Kannen ausgestanzt worden waren, wurde mikroautoradiographisch untersucht. Nach der Applikation von 1–10 mM Chloridlösungen von der der Kannenaußenseite entsprechenden Fläche her waren die Drüsenzellen, die Zellgrenzen im Mesophyll (also die Zellwände einschließlich des dünnen Plasmabelags), Plasmaansammlungen und die Leitbündel besonders stark markiert. Die Drüsenzellen sind dicht mit Plasma erfüllt und wenig vacuolisiert. Die Kornzählungen zeigen, daß die Radioaktivität der verschiedenen plasmareichen Bereiche (der Drüsenzellen, der Zellgrenzen und der Plasmaansammlungen) nicht statistisch signifikant verschieden ist, während die Vacuolen viel weniger markiert sind. Diese Befunde werden im Lichte von Arisz' Theorie des symplasmatischen Transportes diskutiert. Eine artefizielle Übertragung der in den großen Vacuolen der Mesophyllzellen enthaltenen Radioaktivität auf das Cytoplasma während der Präparation kann allerdings nicht völlig ausgeschlossen werden.

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Investigations on the physiology of the glands of carnivorous plantsV. Microautoradiographic investigations of chloride secretion by the gland tissue of Nepenthes

Summary The secretion of ³⁶Cl^- by tissue isolated from Nepenthes pitchers was investigated with microautoradiographic techniques. The tissue was transferred into ether at-30°C or embedded in the vacuum, following procedures described earlier (Lüttge and Weigl, 1965).The microautoradiographs which we obtained after application of 1–10 mM chloride solutions to the surface corresponding to the outside of the pitchers showed a high labelling of the gland cells, the cell borders in the mesophyll (i.e. the cell walls including the thin plasmatic layers), plasmatic particles and the conductive tissue. The gland cells have only very small vacuoles and are densely filled with cytoplasm. The grain counts show that the radioactivity in the parts of the tissue which are rich in cytoplasm (glands, cell borders, plasmatic particles) did not differ within the statistical error. The activity of the vacuoles, however, was much less.These findings are discussed in the light of Arisz' theory of symplasmatic transport. However, an artificial transfer of the radioactivity contained in the large vacuoles of the mesophyll cells to the cytoplasm during the preparation can not be entirely excluded.

     

An ultrastructural and developmental analysis of the corpus allatum of juvenile hormone deficient mutants ofDrosophila melanogaster

Summary The ultrastructure of the corpus allatum of theapterous mutantsap ⁴ andap ^56f ofDrosophila melanogaster during larval-pupal-adult metamorphosis and adult life was correlated with the gland's ability to synthesize juvenile hormone in vitro. During the early wandering period of the third instar of both mutants, a high concentration of smooth endoplasmic reticulum, mitochondria and mitochondrion-scalariform junction complexes are typical features of an active corpus allatum cell. Juvenile hormone biosynthesis by the glands is high at that time and, in fact, only slightly lower than that of wild type glands. In contrast to the wild type gland, the cells of the pupal and pharate adult corpus allatum of both mutants contains highly electron dense mitochondria with tubular cristae but no whorls of smooth endoplasmic reticulum nor glycogen clusters. The frequency and size of the lipid droplets, putatives depots of the juvenile hormone precursors, in cells of theap ^56f gland is a function of the insect's age, but both are lower than in wild type gland cells. Juvenile hormone biosynthesis by both mutant glands remains at the basal level when compared to increased synthesis by the wild type gland. The frequency and density of lipid droplets in cells of theap ⁴ corpus allatum are much lower than in theap ^56f glands. During adult life, the ultrastructural profile of theap ^56f corpus allatum is similar to that of the wild type gland although the in vitro production of juvenile hormone by the former is much lower than that of the wild type gland. The ultrastructural features of the adult corpus allatum ofap ⁴ homozygotes reveal precocious degeneration and support the view that this non-vitellogenic mutant is a juvenile hormone deficient mutation.     

Selection against homozygotes in the ADH polymorphic system of Drosophila melanogaster associated with the lethal factor l(2) Stm

In the natural populations ⁺Tüb, ⁺Prov, and ⁺Rov, similar Adh ^F allele frequencies occur (q _F=0.11, 0.18, and 0.08, respectively). However, there is a discrepancy in that the Adh ^F allele in ⁺Tüb is closely linked to the lethal factor 1(2)Stm, which reduces relative fitness of the F phenotype to zero. In spite of this, polymorphism is maintained also in ⁺Tüb, because the heterozygotes are superior to the homozygous S type (relative fitness=0.88). Under laboratory culture conditions, in ⁺Tüb the relative fitness of the S genotype further decreases to 0.6. After outcrossing the lethal factor, relative fitnesses for S, FS, and F become 0.6, 1, and 0.48, respectively, implying that fitness for S remains the same. Relative values for S, FS, and F in ⁺Prov, not affected by the lethal factor, are calculated by the maximum average fitness method to be 1, 1.2, and 0.2 under the assumption that heterozygous FS are similarly superior to S as in the natural ⁺Tüb population and all allele frequencies found are stable equilibrium values.     

Adventitious bud development and regeneration in Tillandsia eizii

Summary The bromeliad Tillandsia eizii is a stricking species with large, colorful, and persistent inflorescences that can reach 1 m in length. The value of this plant as an ornamental and its importance in cultural and religious activities has led to its overcollection in the wild. Clonal propagation via tissue culture may be a means to repopulate native stands while meeting the demands for this species as an ornamental and ceremonial plant. Adventitious bud proliferation was induced from axenically germinated scedling material. Parameters evaluated were the age of explant material at the time of transfer onto bud-induction medium, the concentration of plant growth regulators, and the period of exposure to induction medium. Light and seanning electron microscopy (SEM) established the origin and development of buds. Twelve-week-old seedling explants rapidly initiated adventitious buds after a 30-d induction period on shoot-initiation medium. Adventitious buds were induced in 40% of the explants placed on media with 2 mg l⁻¹ 6-benzylaminopurine (BA) (8.88 μM) plus 0.1 mg l⁻¹ α-naphthaleneacetic acid (NAA) (0.54 μM) with some cultures becoming highly prolific after repeated subeulture. Shoots elongated in proliferating cultures, and plants were successfully acclimatized and planted into the greenhouse. The results indicate that tissue culture may be used as a means to propagate this epiphytic bromeliad species, which is being seriously affected by deforestation and habitat destruction. In addition, adventitious bud proliferation can provide a means to propagate superior genotypes.     

Precise temporal regulation of roughest is required for correct salivary gland autophagic cell death in Drosophila

The Drosophila roughest (rst) locus encodes an immunoglobulin superfamily transmembrane glycoprotein implicated in a variety of embryonic and postembryonic developmental processes. Here we demonstrate a previously unnoticed role for this gene in the autophagic elimination of larval salivary glands during early pupal stages by showing that overexpression of the Rst protein ectodomain in early pupa leads to persistence of salivary glands up to at least 12 hours after head eversion, although with variable penetrance. The same phenotype is observed in individuals carrying the dominant regulatory allele rst^D, but not in loss of function alleles. Analysis of persistent glands at the ultrastructural level showed that programmed cell death starts at the right time but is arrested at an early stage of the process. Finally we describe the expression pattern and intracellular distribution of Rst in wild type and rst^D mutants, showing that its downregulation in salivary glands at the beginning of pupal stage is an important factor in the correct implementation of the autophagic program of this tissue in space and time. genesis 47:492–504, 2009. © 2009 Wiley‐Liss, Inc.     

Molecular Cloning and Characterization of a Vacuolar H+-pyrophos-phatase Gene, SsVP, from the Halophyte Suaeda salsa and its Overexpression Increases Salt and Drought Tolerance of Arabidopsis

The chenopodiaceae Suaeda salsa L. is a leaf succulent euhalophyte. Shoots of the S. salsa are larger and more succulent when grown in highly saline environments. This increased growth and water uptake has been correlated with a large and specific cellular accumulation of sodium. S. salsa does not have salt glands or salt bladders on its leaves. Thus, this plant must compartmentalize the toxic Na⁺ in the vacuoles. The ability to compartmentalize sodium may result from a stimulation of the proton pumps that provide the driving force for increased sodium transport into the vacuole. In this work, we isolated the cDNA of the vacuolar membrane proton-translocating inorganic pyrophosphatase (H⁺-PPase) from S. salsa. The SsVP cDNA contains an uninterrupted open reading frame of 2292 bp, coding for a polypeptide of 764 amino acids. Northern blotting analysis showed that SsVP was induced in salinity treated leaves. The activities of both the V-ATPase and the V-PPase in Arabidopsis overexpressing SsVP-2 is higher markedly than in wild-type plant under 200 mM NaCl and drought stresses. The Overexpression of SsVP can increase salt and drought tolerance of transgenic Arabidopsis. Electronic supplementary material Electronic supplementary material is available for this article at and accessible for authorised users. Shanli Guo, Haibo Yib: These authors contributed equally to this work     

The effects of the glucocorticoid,dexamethasone, on the development of the Drosophila embryo

We have investigated the effects of the glucocorticoid, dexamethasone, and five structural analogs on Drosophila development in an effort to identify steroid ligands that may play a role in the embryogenesis of this organism. Embryos were exposed to glucocorticoids either by direct culture in supplemented medium, or by examining embryos from adult flies raised on supplemented fly food. After exposure, embryos were examined for developmental defects. At a morphological level, exposure to dexamethasone disrupts the dorsolateral folding of the amnioserosa during germ band extension. In addition, germ band retraction and dorsal closure is also disrupted. The phenocritical period of these effects is within the first 4 h of embryogenesis. This response is dosage sensitive, with embryos responding to concentrations of dexamethasone ranging from 10^–6 to 10^–3M. Furthermore, glucocorticoids which are closely related structural analogs of dexamethasone also disrupt germ band retraction and dorsal closure, while other tested steroids had no effect on embryonic development. At a molecular level, expression of the gene, Krüppel, is absent from the amnioserosa of dexamethasone-treated embryos. The cuticular phenocopy resulting from exposure to dexamethasone and related glucocorticoids is morphologically similar to the mutant phenotype associated with four genes required for germ band retraction, namely hindsight, serpent, tail-up and u-shaped. The results of this study represent the first association of a glucocorticoid with dose, stage and tissue specific effects on Drosophila development at both morphological and molecular levels.     

Autonomy of the wingless mutation in Drosophila melanogaster

Summary T(Y;2) translocations were used to cytologically localise the wingless locus of Drosophila melanogaster. We found that an existing T(Y;2), which is an insertion of a segment of 2L into the Y chromosome, has wg ⁺ within this insert. This Y chromosome was used to generate an attached XY chromosome containing wg ⁺. The mutation claret-nondisjunctional (ca ^nd) was used to induce the loss of this XY chromosome and thus generate gynandromorphs with wg ¹/wg ¹ male tissue and wg ⁺/wg ¹/wg ¹ female tissue. Analysis of these gynanders demonstrated that a genotypically wingless mutant hemithorax is usually also phenotypically mutant in these half body mosaics; thus wg ¹ is discautonomous. This observation is of interest as it is known that wg is not cell autonomous.     

Coral-crab association: a compact domain of a multilevel trophic system

Colonies of the Red Sea branching coral Stylophora pistillata were incubated in situ with radioactive carbon and Trapezia cymodoce crabs were introduced to the colonies, for one month each, up to 7 months after coral labelling. Zooxanthellar photosynthetic products were translocated to the crabs via host coral tissue. Based upon crab/coral tissue conversion from 53 crab samples, crabs possessed radioactive material equal to that recorded in 320–770 mm² of coral tissue (up to 2257 mm²). This material was translocated mainly by direct grazing on coral tissue rather than mucus collection. Ovigerous female crabs (39 specimens) accumulated significantly more labelled carbon than males, and up to 53 % of their radioactivity was concentrated in their reproductive organs. A pair of crabs dwelling in a coral colony consumed ca 130 cm² of host tissue per month (40–45 cm length of coral branches). This system represents a compact, obligatory multilevel trophic domain which also radiates horizontally by allocation of energy derived from algal photosynthesis into planula and zooea larva production, permitting the development of long trophic chains and a complex food web.     

Reinvestigation of auxin and fusicoccin stimulation of the plasma-membrane H+-ATPase activity

In vivo treatment of maize (Zea mays L.) coleoptile segments with auxin (indole-3-acetic acid; IAA) and fusicoccin (FC) followed by plasma-membrane isolation was used to characterize the effects of these treatments on the plasma-membrane H⁺-ATPase. Both IAA and FC increased H⁺ extrusion and elongation rate of the coleoptile segments, FC more strongly than IAA. Plasma membranes isolated after in-vivo treatment with FC showed a twofold stimulation of ATP hydrolysis and a several-fold stimulation of H⁺ pumping, whereas no effect was observed after IAA treatment, irrespective of whether the plasma membranes were prepared by two-phase partitioning or sucrose-gradient centrifugation. A more detailed investigation of the kinetic properties and pH dependence of the enzyme showed that FC treatment led to a twofold increase in V _max, a decrease in K _m for ATP from 1.5 mM to 0.24 mM, and a change in pH dependence resulting in increased activity at physiological pH levels. Again, IAA treatment showed no effects. Quantitation of the H⁺-ATPase by immunostaining using four different antibodies revealed no difference between IAA-and FC-treated material, and controls. From these data we conclude that (i) neither IAA nor FC gives rise to an increase in the amount of H⁺ -ATPase molecules in the plasma membrane that can be detected after membrane isolation, and (ii) if the H⁺-ATPase is activated by IAA, this activation is, in contrast to FC activation, not detectable after membrane isolation.Abbreviations BTP 1,3-bis(tris[hydroxymethyl]methylamino)-propane - FC fusicoccin - lyso-PC lysophosphatidylcholine - Mes 2-(N-morpholino)ethanesulfonic acid This paper is dedicated to Prof. Dieter Klämbt on the occasion of his 65th birthdayWe thank Ann-Christine Holmström and Adine Karlsson for excellent technical assistance, Professor Ramón Serrano (Instituto de Biologia Molecular y Celular de Plantas, UPV-CSIC, Universidad Politecnica, Valencia, Spain) for a generous gift of antisera to the H⁺-ATPase and Professor Wolfgang Michalke (Institut für Biologie III, Albert-Ludwigs-Universität, Freiburg, Germany) for kindly providing the monoclonal antibody to the H⁺-ATPase. This work was supported by the Swedish Natural Science Research Council, the Deutsche Agentur für Raumfahrtangelegenheiten (DARA, Bonn) via AGRAVIS (Bonn) and by the Ministerium für Wissenschaft und Forschung (MWF, Düsseldorf). Thomas Jahn received scholarships from the Deutsche Graduiertenförderung des Landes Nordrhein-Westfalen and the Deutscher Akademischer Austauschdienst (DAAD, Bonn).     

Coal Dust Pollution Effects on Wetland Tree Species in Richards Bay,South Africa

In this study, the effects of coal dust on four, sympatric, wetland tree species in Richards Bay Harbour were investigated. We tested the hypothesis that leaf micromorphology influenced dust accumulation and that coal dust occluded stomata and reduced photosynthetic performance of three mangroves, Avicennia marina, Bruguiera gymnorrhiza and Rhizophora mucronata, and a mangrove associate, Hibiscus tiliaceus. To investigate leaf micromorphology, leaf blade material of the four species was prepared following standard procedures and viewed under scanning electron microscopy. Gas exchange and chlorophyll fluorescence measurements were made at saturating light (>1000 μmol m⁻² s⁻¹) and high temperature (>25 °C) on leaves that were either covered or uncovered with coal dust. There was no evidence of occlusion of stomata by dust. Dust accumulation in A. marina and H. tiliaceus was exacerbated by the presence of a dense mat of trichomes on the undersurface of the leaves, as well as by the sticky brine secreted by salt glands in the former species.Coal dust significantly reduced CO₂ exchange, Photosystem II (PS II) quantum yield and electron transport rate (ETR) through PS II in A. marina and H. tiliaceus but not in the other two mangroves. Reduction in photosynthetic performance was attributed to reduction in light energy incident on the photosynthetic tissues.     

Stabilität des Phagen PS8 in Crown-Gall-Gewebekulturen von Vinca rosea L.

Zusammenfassung Aus einem A. tumefaciens, B6, infizierten Sproß von Vinca rosea wurde eine Tumorgewebekultur hergestellt, die in 2 verschiedenen Tests bakterienfrei war. Beim Plattieren von Homogenaten dieser Kultur gegen den phagensensitiven Stamm B6-806 von A. tumefaciens wurden keine Phagen gefunden. Auch Hitzeschock, Bestrahlung mit UV oder Behandlung mit Mitomycin induzierten keinen Phagen. Das Homogenat selbst inaktiviert zugefügten Phagen PS8 nicht.Bei 26° C verliert PS8 seine Aktivität in 13 Wochen (von 10⁷PS8/ml auf 0). Wenn PS8 einer wachsenden Tumor-Gewebekultur zugefügt wird, verliert er rascher an Aktivität. Dies ist z. T. Folge der Verdünnung des Phagen durch die wachsende Gewebekultur, z. T. Folge einer Wechselwirkung zwischen Phage und Gewebe. In der Tumorkultur wurde keine Vermehrung des Phagen PS8 beobachtet.Die Ergebnisse werden im Hinblick auf tumorigene Eigenschaften von PS8 diskutiert.

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Stability of phage PS8 in crown-gall tissue cultures of Vinca rosea L.

From Vinca rosea shoots a tumor tissue culture (Agrobacterium tumefaciens B6) was prepared which was bacteria-free in 2 experimental tests. This culture did not contain any phages when homogenates of the tissue were plated against the phage-sensitive strain B6-806 of Agrobacterium tumefaciens. Also heat shock, UV irradiation or treatment with mitomycin C did not induce any phage.The homogenate itself did not interfere with the activity of added phage PS8. At room temperature the bacteriophage PS8 looses activity within 13 weeks (10⁷ PS8/ml to 0).If PS8 is added to a growing tumor tissue culture the loss of activity is more rapid, partly because the phage is diluted by the growing culture, partly because some interaction between tissue culture and phage activity. No propagation of PS8 in the tumor tissue culture was observed.These results are discussed in relation to the hypothesis of PS8 as the tumor inducing principle.

     

Assessment of Cell Viability in Intact Glandular Tissue in <Emphasis Type="Italic">Chironomus ramosus</Emphasis> using Dye-exclusion and Colorimetric Assays

Conventionally, dye-exclusion test for determining cell viability has been restricted only for cells in suspension in tissue culture. In this paper, salivary gland of Chironomus has been proposed as a simple tissue model system where dye-exclusion test can be reliably employed for the intact gland. We have compared suitability of commonly used vital dyes and nigrosin was found suitable for the salivary gland cells. Biochemical tests using tetrazolium salts are also commonly used for determining quantitative indices of cell viability in metabolically active cells. Ours is the first attempt to extend the same technique for the whole tissue. We standardized the conditions and prepared a protocol for MTT-based colorimetric assay suitable for the salivary gland of Chironomus. A strong correlation (r² = 0.9893) was obtained where increasing O.D. correlated linearly with the number of live glands. We concluded that nigrosin dye-exclusion and MTT metabolic inclusion assays are suitable methods for the viability test of metabolically active intact salivary gland of Chironomus which can serve as a potential model for the assessment of cytotoxicity in future.     

Nachweis von häutungsaktiven Stoffen in isolierten Prothorakaldrüsen und Oenocyten beiBombyx mori während des 5. Larvenstadiums

Summary The content of moulting hormones has been determined in homogenates of isolated prothoracic glands and oenocytes during the 5th instar of the silkworm,Bombyx mori by means of the Calliphora bioassay.Prothoracic glands show variable activity in the production of moulting hormones, reaching a maximum near the end of the larval period. Comparable activities, but at higher levels, could be demonstrated in oenocytes. Controls with doubled quantities of tissue produced in a proportionate reaction in the bioassay. Fat bodies were inactive.Prothoracic glands and oenocytes incubated together resulted in a slower pupation index than would be expected from the sum of single determinations of oenocytes and prothoracic glands. This is explained by the ability of prothoracic glands to build conjugates of ecdysones.Die Arbeit wurde mit Unterstützung der Deutschen Forschungsgemeinschaft durchgeführt     

Plant cell graft chimeras obtained by co-culture of isolated protoplasts

Summary Heterospecific chimeralSolanum nigrum (+)Solanum tuberosum plants were obtained by cell grafting in protoplast co-cultures. Periclinal, sectorial, and mericlinal chimeras have been identified by various morphological and cytological characteristics.Morphogenesis predominantly began in periclinal chimeral organization. Cells of different species have been found to be interconnected by secondary plasmodesmata. Plantlets of all chimeral lines were grown to flowering under tissue culture conditions and some also in the greenhouse. Aspects of organogenesis and interspecific cooperation are discussed.Abbreviations B 5 culture medium (Gamborg et al. 1968) supplemented with 2.5M 6-benzyladenin - L1, L2, L3 epidermis (L1), subepidermal layer (L2), core (L3) - MS culture medium (Murashige andSkoog 1962) - n, n _w symbols used for the indication ofSn-F (n) orSn-F-W2 (n _w ) tissue in L1, L2 or L3 - Sn Solanum nigrum - Sn-F is an atrazine-resistant biotype - Sn-F-W2 is a plastid mutant malbino derivative ofSn-F - St Solanum tuberosum - St-H²258 is a dihaploid clone - t symbol used the indication ofSt-H²258 tissue in L1, L2 or L3 - V-KM culture medium (Binding and Nehls 1977) Dedicated to Professor Dr.Josef Straub, late Director of the Max-Planck-Institute für Züchtungsforschung at Cologne, who was the first to study the production of chimeras by callus association in 1972.     

Filamentous algae in fish ponds of the Třeboň Biosphere Reserve-ecophysiological study

Filamentous algae in eutrophic carp ponds in South Bohemia (Central Europe) were studied from 1988 to 1990. High biomass (490 g DW m^-2) was attained by Cladophora fracta (O. F. Müll. ex Vahl) Kütz. after two months of growth. This marked growth depleted inorganic carbon in the water, but did not decrease the concentration of tissue nutrients. Laboratory measurements of final pH indicate that all the filamentous algae studied, except for Tribonema, are very efficient HCO₃ ^- users. An extremely high pH of 11.6 and oxygen concentration of 32 mg l^-1 were measured in the algal mats. High pH resulted in CaCO₃ precipitation, visible as white incrustations on algal filaments. The amount of precipitated CaCO₃ reached 134 kg ha^-1. After reaching peak biomass, 90% of the Cladophora decomposed over the next 95 days.The highest net photosynthetic rate in C. fracta was measured between pH range 8.5–10.0 and oxygen concentrations of 7–12 mg l^-1. Optimum temperature for photosynthesis was between 17–22°C.     

Laser microdissection and cryogenic nuclear magnetic resonance spectroscopy: an alliance for cell type-specific metabolite profiling

Laser microdissection was used as a tool to harvest secretory cavities (SC) from leaves of Dilatris pillansii Barker (Haemodoraceae) and from leaves and flowers of herbarium specimens of Dilatris corymbosa Berg. and Dilatris viscosa L. Cryogenic ¹H NMR spectroscopy and HPLC analysis of microdissected samples indicated specific accumulation of methoxyphenylphenalenones in the SC. The structures of two novel and a known natural product in the secretory tissue were confirmed by comparison with authentic compounds isolated from rhizomes and roots from which further phenylphenalenones and phenylphenalenone glucosides were isolated and identified by spectroscopic methods. How it will be possible to use the LMD technique to localize natural products in specific plant cell populations is also discussed. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users. An erratum to this article can be found at     

Die Speicheldrüsen junger Ratten nach Parasympathektomie

Zusammenfassung Bei 13 und 28 Tage alten Ratten, deren Speicheldrüsen noch undifferenziert sind, wurde die Chorda tympani in der Paukenhöhle durchtrennt, 1–123 Tage nach der Operation wurden die Tiere getötet und Gl. submandibularis und Gl. sublingualis untersucht. Zu Beginn der Arbeit wird die postnatale Entwicklung dieser Drüsen beschrieben.Nach dem Eingriff läßt sich in der Gl. submandibularis eine Beschleunigung der postnatalen Entwicklung feststellen, als deren Ursache die herabgesetzte Funktionsaktivität der Drüsenzellen angesehen wird.In der Gl. sublingualis treten stellenweise Nekrosen von Azinusgruppen und ausgeprägte Veränderungen an den Ausführungsgängen auf, die den Befunden bei experimenteller Hydrodyschylie (Seifert, 1964) ähneln. Außerdem findet sich an beiden Drüsen nach einer initialen Sekretanhäufung in den Azinuszellen eine deutliche Zellatrophie (in Verbindung mit Hypotrophie und Hypoplasie?) mit Gewichtsverlust der Drüsen bis maximal 51%. An diese Atrophie schließt sich eine teilweise sich abspielende Restitution der Drüsen an. Die Befunde werden mit denen verglichen, die an den Speicheldrüsen erwachsener Ratten nach Parasympathektomie erhoben wurden (Müller, 1967).

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Salivary glands of young rats following parasympathectomyMorphological and histochemical studies of the submandibular and major sublingual salivary glands

Summary The left chorda tympani nerve was cut in the cavum tympani of 13 and 28 days old albino rats. The animals were killed at intervals over a period from 1–123 days following the operation. A survey of the postnatal development of the submandibular and sublingual glands is given.In the denervated submandibular gland there was evidence of an accelerated postnatal development. It is assumed that the decreased functional activity of the gland is the cause of this acceleration.The sublingual gland of the operated side showed areas of necrotic acini and alterations of the ducts similar to those found at experimental hydrodyschylia (Seifert, 1964).Besides both glands show an initial increase of secretion products within the cells of the acini which is followed by an atrophy of the glands with reduction in size and loss of weight (Maximum 51%). This atrophy is followed by partial restitution of the glands. The findings are compared with those found in denervated glands of adult rats (Müller, 1967).

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Herrn Professor Dr. Kurt Goerttler zum 70. Geburtstag gewidmet.

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Mit dankenswerter Unterstützung durch die Deutsche Forschungsgemeinschaft.