Evidence for calcium translocation in catch connective tissue of the sea cucumber Stichopus chloronotus

Summary Catch connective tissue is collagenous connective tissue that changes its mechanical properties rapidly. Calcium has been assumed to play a key role in the catch mechanism because calcium concentration profoundly affects the mechanical properties. We have found a calcium-storage cell, the vacuole cell, in the dermis of the sea cucumber Stichopus chloronotus a wellstudied catch connective tissue. The processes of this cell are packed with vacuoles measuring 0.1–1.5 m in diameter. The ultrastructure was compared in a relaxed state and in a catch (stiff) state induced by high-potassium artificial sea water. When fixed with pyroantimonate, the vacuoles contained precipitates in the relaxed state but not in the catch state. Analysis with an energy-dispersive X-ray microanalyzer suggested that the precipitate contains calcium. The release of calcium ions from the vacuole cell is proposed to induce connective tissue catch.     

Ultrastructure of the flagellar apparatus in the green flagellateSpermatozopsis similis

The ultrastructure of the flagellar apparatus of the naked, biflagellate green algaSpermatozopsis similis Preisig & Melkonian has been studied in detail using an absolute configuration analysis. The two basal bodies are displaced by 350 nm in the 1/7 o'clock direction and do not overlap proximally. They are interconnected by a principal distal connecting fibre consisting of a bundle of 5–8 nm filaments and possibly two proximal striated connecting fibres. The flagellar root system is cruciate (5-2-5-2 or 4-2-4-2 system) and contains a prominent continuous system I fibre overlying the two opposite two-stranded roots. A system II fibre is absent. Pronounced structural differences have been observed in the flagellar apparatus ultrastructure at two types of flagella orientation: During backward swimming basal bodies are parallel, the distal connecting fibre is extremely contracted; during forward swimming basal bodies assume various angles (from 20° to 180°) and the connecting fibre is about five times longer compared to the contracted state. The function of the connecting fibre as a contractile organelle and the mechanism of its contraction are discussed. On the basis of the flagellar apparatus ultrastructure,Spermatozopsis similis is related toChlamydomonas-type green algae.     

The ultrastructure of the sinus gland of Carcinus maenas (Crustacea: Decapoda)

Summary The sinus gland of Carcinus maenas consists of the swollen axonal endings of the neurosecretory cells of the major ganglia and acts as a storage release centre for the membrane bound neurosecretory material. These neurosecretory granules fall into five different types based on size and electron density. Their contents are released by exocytosis of the primary granules or smaller units budded from the primary granules.I thank Professor E. Naylor for his constant advice and Professor E. W. Knight-Jones, Department of Zoology, University College, Swansea, for the provision of laboratory facilities. I am grateful to the Science Research Council for the financial support. Finally, I thank the Electron Microscope Unit, Southampton General Hospital, where the work was completed.     

Fine structural and cytochemical study of the innervation of smooth muscle in an amphibian (Bufo marinus) lung before and after denervation

Summary The innervation of the toad (Bufo marinus) lung was studied with transmission electron microscopy and fluorescence techniques, both before and after 12 or 20 days close vagosympathetic denervation. Four cytologically distinct types of neuronal processes were recognised, in relation to the visceral muscles of the lung. These were described as cholinergic, adrenergic, nonadrenergic/non-cholinergic (NANC) and sensory on the basis of the characteristics of their vesicular content and cytochemical reactions. An apparent efferent innervation of visceral smooth muscle was achieved by NANC (50%), cholinergic (25%) and adrenergic (25%) fibres. A few sensory fibres were also present. After denervation only NANC fibres persisted, showing that the cell bodies of these fibres were intrapulmonary. The vascular smooth muscle was supplied by cholinergic, adrenergic and sensory fibres. In the walls of the proximal branches of the pulmonary artery were fibres containing large dense-cored vesicles. These profiles, which were associated with the vasa vasorum, were similar to neurosecretory fibres. After denervation all neural profiles associated with the vasculature had degenerated. The observations suggest that vagal vasodepressor effects in the toad lung are mediated indirectly through relaxation of visceral muscle strands which in their contracted state compress vascular channels.The authors would like to thank Dr. J.R. McLean for technical advice on fluorescence microscopy. This work was supported by a grant from the Australian Research Grants Committee     

Length of burning rotation and community composition in high-level Calluna-Eriophorum bog in N England

Studies on a long-term experiment on Calluna-Eriophorum bog indicate that the length of time between management fires affects the post-fire development. A short rotation (every 10 years) results in increased dominance by Eriophorum spp. while a long rotation (every 20 years) results in greater abundance of Calluna vulgaris after fire. Variations in post-fires species abundances are related to pre-fire stand composition coupled with the patchy distribution of regeneration centers of rhizomatous species. The usefulness of burning this vegetation type in management for herbivore grazing is questioned.Nomenclature follows Tutin et al. (1964 80) for vascular plants. Smith (1978) for mosses, Watson (1981) for liverworts and Hawksworth, James & Coppins (1980) for lichens. Following common practice. Calluna vulgaris is referred to simply as Calluna. Acknowledgements. I thank M. Raves and the Nature Conservancy Council for permission to work on Moor House National Nature Reserve and for the use of data from Reserve Records, V. J. Hobbs for field assistance, and Prof. C. H. Gimingham for help and advice during the study. I also thank all who worked at Moor House for creating a pleasant working environment. The long-term experiment was planned by Dr R. J. Elliot and analyzed initially by Dr G. I. Forrest.     

Accumulation of UV-absorbing flavonoids induced by UV-B radiation inAmbidopsis thaliana L.

Irradiation ofArabidopsis with ultraviolet (UV) light resulted in intensity- and wavelength-dependent increases in the levels of a small family of UV-absorbing flavonoids, which accumulate in the aerial parts of the plants. A gradient of sensitivity to UV-B radiation is described in the different leaves of developingArabidopsis plants whereby the earliest formed leaves become damaged by UV-B faster and more extensively than later formed leaves. This UV-sensitivity gradient tightly parallels differences in constitutive as well as UV-induced levels of flavonoid accumulation among the various leaves, suggesting a direct role of flavonoids in protection against damage by UV radiation. The level of accumulated flavonoids, both constitutive and UV-induced, in each leaf appear to be dependent on the specific developmental state of each leaf as well as the overall developmental state of the plant. The UV-mediated flavonoid response, along with the observed UV-induced damage, appear not to be systemic inArabidopsis but restricted very closely to the irradiated areas of leaves.I am deeply indebted to Robert Fischer and Bob Buchanan for providing access to their laboratories and for their invaluable help without which this work could not have been realized. I especially wish to thank Lola Peñarrubia, Elena del Campillo, Patrick Neil and Julie Montgomery for innumerable and fruitful discussions. This work was supported by Cooperative State Research Service, U.S. Department of Agriculture, under Agreements Nos. 90-37280-5664 and 90-372780-5808.     

Field observations on nitrogen catch crops. III. Transfer of nitrogen to the succeeding main crop

In temperate climates with a precipitation surplus during autumn and winter, nitrogen (N) catch crops can help to reduce nitrogen losses from cropping systems by absorbing nitrogen from the soil and transfer it to a following main crop. In two field experiments the catch crop species winter rye (Secale cereale) and forage rape (Brassica napus ssp. oleifera (Metzg.) Sinsk) or oil radish (Raphanus sativus spp. oleiferus (DC.) Metzg.) were planted end of August and 3 weeks later with a non-limiting supply of N and zero-N controls. In the next spring catch crops were incorporated into the soil. In Expt 1, N transfer was measured as (i) the N uptake of a potato test crop, grown with zero and 12.5 g m^–2 N applied, and (ii) the increase in soil mineral N (0–30 cm) in uncropped soil covered with polythene film. In Expt 2, N transfer was measured as the increase in soil mineral N in covered cylinders placed in uncropped soil (in situ incubation). Subsidiary laboratory incubations were performed in Expt 2. In Expt 1, the apparent recovery in potato of fertilizer N (R _f) was 0.56. The recovery in potato of N mineralized from 'native' N pools other than catch crop material (R _n) ranged from 0.43 to 0.51, depending on the value assumed for the depth of N extraction by potato roots. The average recovery in potato of incorporated catch crop N (R _c) was 0.34. Expressed as `fertilizer N replacement factor' (F _r) the latter was 0.61 (i.e. 1 kg of N in catch crop material counts for 0.61 kg fertilizer N). Under the film in Expt 1 the fraction net mineralization of incorporated catch crop N (M _n) was 0.36 on August 11 and 0.43 on October 18. In Expt 2, the average value of M _n was 0.31, which was lower than in Expt 1 and probably associated with the drier soil in Expt 2. In the laboratory incubations (20°C) M _n showed values up to 0.54 after 84 days with the largest rates of change in mineralization occuring early after the start of the incubation. In conjunction with literature data it is concluded that cultivation of nitrogen catch crops shows promise as a means to reduce N input and N losses in temperate climates with wet winters.     

Catch Composition on Red Shrimps’ (Aristaeomorpha foliacea and Aristeus antennatus) Grounds in the Eastern Ionian Sea

In the present study, the catch composition and the catch per unit effort (CPUE) by weight and numbers in red shrimps’ (Aristaeomorpha foliacea and Aristeus antennatus) grounds was examined in the southern part of the eastern Ionian Sea, in order to collect important information for the Greek waters, where no deep-water fishery exists. In the depth stratum 500–700 m, the catch of the commercial species represented a high proportion (>70%) of the total catch. Red shrimps and several other commercial species were found in important quantities. The present results suggest the possibility of developing a deep-water fishery in Greece. In such a case, attention should be paid because of the high vulnerability of A. foliacea – the main deep-water fishing resource in the area – to the fishing pressure.     

Ultrastructure of the neurogenic heart of Limulus polyphemus

Summary The ultrastructure of Limulus cardiac muscle was examined. The hearts were fixed in situ by perfusion with isotonic glutaraldehyde solution while in relaxed, contracted, or stretched states. The sarcomeres are relatively long, varying in length from about 2.5 to 6.6 . The average A-band length is 2.46 . M lines are absent, and H zones are poorly distinguished. Thick and thin filament diameters average about 200 Å and 50 Å, respectively; each thick filament is surrounded by 8–12 thin ones. Superficial invaginations of the sarcolemma occur, making contact with the Z lines of the outermost myofibrils. There is an extensive sarcoplasmic reticulum and transverse (T) tubules. Some T tubules run longitudinally and some open into deep sarcolemmal invaginations which extend into the fiber interior. The T tubules swell markedly in hypertonic solution. Single neurons and small bundles of neurons are observed in close apposition with myocardial cells. Intercalated disks are found in Limulus heart at regions of contact between contiguous myocardial cells lying end to end; semitight or gap junctions are essentially absent. Prominent differences in sarcomere lengths sometimes occur across the disk, thus indicating that the disks demarcate cells functionally. Hence, in addition to direct motoneuron activation, there may be some transfer of excitation across the intercalated disks in accord with our previous finding that propagating, overshooting action potentials can be induced in this heart.Supported by grants from the American Heart Association and from the Public Health Service (HE-11155 and HE-05815). I thank Mrs. Jan Redick for expert technical assistance.     

Apical surface of the epithelial cells in the gallbladder of the rainbow trout and the tench

Summary The aluminium-formaldehyde (ALFA) histofluorescence method was used to study the innervation of the gill of the marine bivalve mollusc Mytilus edulis and the results were contrasted with those obtained with the standard formaldehyde-induced-fluorescence (FIF) method. The ALFA method produced more fluorescing structures than the FIF method, thus revealing fine branches of the branchial nerve running beneath the gill epithelium which previously remained undetected. This study demonstrates the usefulness of the ALFA histofluorescence method in the study of marine invertebrates.This study was supported in part by Grants 1506RR08171 from NIMH and 5T32GM07641 from the MARC Program of NIGMS. I wish to thank E. Aiello for thoughtful discussions of the work     

Further observations of transport within paddle cilia

Summary Rhabdopleura has densely ciliated tentacles and cephalic shield. These cilia are of two sorts, ordinary cilia and those bearing paddle-like swellings. These swellings, about l m across, are in the ciliary membrane. The axoneme bundle within the cilium remains intact, keeping the usual spatial arrangements, except that in the swelling its overall diameter is reduced. It is suggested that the mechanism for moving the paddle-like swellings along the length of the cilium is the releasing and reattaching of the side filaments from the axoneme to the ciliary membrane. Acknowledgements. I wish to thank the Director and staff of the Plymouth Marine Biological Association for their help and facilities while collecting the material. Mr. R.F. Moss gave much skilled technical assistance. I wish also to thank Professor W.E. Knight-Jones who identified the specimens of Vermiliopsis and Dodecacera.     

The bacterial oxidation of nicotinic acid

N-Formylmaleamic acid, a probable intermediate in the bacterial metabolism of nicotinic acid, has been synthesized by photoisomerization of its transisomer, N-formylfumaramic acid. The compound previously reported to be N-formylmaleamic acid has been shown to be N-formylfumaramic acid.This paper is dedicated to Prof. R. Y. Stanier, who served as a model of elegant erudition to me during my graduate work with him.I thank the NIH and NSF for support     

On the ultrastructure of polypeptide hormone-producing cells in the gut of the ascidian,Ciona intestinalis L. and the Bivalve,Mytilus edulis L.

Summary Ultrastructural evidence has been found for the presence of polypeptide hormone-producing cells in the gut of Ciona intestinalis L. and Mytilus edulis L. which do not appear to have been described before. Due to their localization and ultrastructural characteristics, it is suggested that the cells in Mytilus edulis probably produce an insulin-like substance and that some of these cells in Ciona intestinalis may produce 5-HT (5-Hydroxytryptamine). In each species only one granulated cell type can be observed. The granules, which are electron dense and membrane bound, also show a halo. The average diameter of the granules is 100–200 nm for Ciona and 200–400 nm for Mytilus.I thank Mr. G. Bargsten, M.A., Dept. of Marine Zoology, University of Kiel, for the supply of the animals     

Extension-growth responses and expression of flavonoid biosynthesis genes in the Arabidopsis hy4 mutant

The hy4 mutant of Arabidopsis thaliana(L.) Heynh. was previously shown to be impaired in the suppression of hypocotyl extension specifically by blue light. We report here that hy4 is altered in a range of blue-light-mediated extension-growth responses in various organs in seedlings and mature plants: it shows greater length of bolted stems, increased petiole extension and increased leaf width and area in blue light compared to the wild type. The hy4 mutant shows decreased cotyledon expansion in both red and blue light compared to the wild type. Anthocyanin formation and the expression of several flavonoid biosynthesis genes is stimulated by blue light in the wild type but to a much lower extent in hy4. The results indicate that the HY4 gene product is concerned with the perception of blue light in a range of extension-growth and gene-expression responses in Arabidopsis.Abbreviations DFR dihydroflavonol reductase - CHS chalcone synthase - CHI chalcone isomerase We thank the UK Agricultural and Food Research Council for supporting this work through the award of a research grant to G.I.J. We are grateful to Robert Brown for excellent technical assistance and Drs B.W. Shirley (Department of Biology, Virginia Polytechnic Institute and State University, Blacksburg, USA), C.D. Silflow (Department of Genetics and Cell Biology, University of Minnesota, St. Paul, USA) and I.E. Somssich (Department of Biochemistry, Max-Planck-Institut, Köln, Germany) for providing plasmid DNA.     

Lipid transfer protein genes specifically expressed in barley leaves and coleoptiles

Genes/cDNAs encoding so-called lipid-transfer proteins (LTPs) have been isolated from a variety of tissues from different plants, but the in-vivo function of the LTP proteins is not yet known. In barley (Hordeum vulgare L.), the LTP1 gene (encoding a probable amylase/ protease inhibitor, Mundy and Rogers 1986, Planta 169, 51–63) is active in aleurone tissue, and in this paper two LTP-encoding cDNAs isolated from green leaves are described. The encoded proteins start with signal sequences, they are 75% homologous to each other, 60–63% homologous to rice aleurone LTP and maize seed/ coleoptile LTP, but only 48% homologous to barley aleurone LTP. Northern hybridization experiments established that the two seedling-specific genes are both highly expressed in leaves and coleoptiles whereas the LTP1 gene is inactive in seedlings. No LTP gene expression was detected in roots using either seedling or aleurone cDNA clones as probes. Tissue-print hybridization indicates that the LTP genes are first expressed in young epidermal cells in leaves and coleoptiles, and subsequently expressed in the vascular strands. Genomic Southern analysis indicates that the barley LTP gene family has four to six members.Abbreviation LTP lipid transfer protein I thank Dr. J. Mundy, Carlsberg Research Laboratory, Copenhagen, Denmark for the PAPI cDNA clone and R. Barkardottir, Department of Molceular Biology, University of Aarhus, Denmark for providing RNA for some of the Northern analyses. I also thank I. Bjørndal and L. Kjeldbjerg for excellent technical assistance. This work was supported by the The Danish Biotechnology Programme.     

Interactions within and between species at different densities of the bark beetleIps typographus and its predatorThanasimus formicarius

Interactions between the bark beetleIps typographus and one of its predators,Thanasimus formicarius, were investigated in caged spruce logs containing both species in eight different density combinations. Productively (offspring per female) ofIps was adversely affected by highIps gallery density as well as highThanasimus density.Thanasimus productivity was enhanced by highIps gallery density but negatively affected by highThanasimus density.Ips productivity andThanasimus developmental rate differed between tree individuals, probably owing to tree-related differences in phloem thickness. Relative predator-causedIps mortality was ca 20% higher at high gallery density (ca 300 egg galeries per m²) than at low gallery density (ca 100 egg galleries per m²), indicating that mortality was density dependent. This difference was due to the fact thatThanasimus larval density was positively related toIps gallery density. Mortality increased by ca 0.4% with each additionalThanasimus larva per m², independently of gallery density. Relative population levels ofIps andThanasimus were monitored with pheromone traps in two regions differing in theirIps typographus outbreak history. Absolute catch as well as theThanasimus/Ips catch ratio were ten times greater in the outbreak region than in the non-outbreak region. Coupled with the results in the caging experiment, this indicates thatT. formicarius responds numerically to changes inI. typographus numbers per unit bark area as well as toIps population changes at the regional level. The findings suggest that predation under bark may be a significant factor in supressingI. typographus outbreaks.     

Effects of temperature on the activity of phosphoenolpyruvate carboxylase and on the control of CO2 fixation in Bryophyllum fedtschenkoi

The phosphorylation state and the malate sensitivity of phosphoenolpyruvate carboxylase (PEPCase, EC 4.1.1.31) in Bryophyllum fedtschenkoi Hamet et Perrier are altered by changes in the ambient temperature. These effects, in turn alter the in-vivo activity of the enzyme. Low temperature (3 °C or less), stabilizes the phosphorylated form of the enzyme, while high temperature (30 °C) promotes its dephosphorylation. The catalytic activity of the phosphorylated and dephosphorylated forms of PEPCase increases with temperature, but the apparent K _i values for malate of both forms of the enzyme decrease. Results of experiments with detached leaves maintained in darkness in normal air indicate that the changes in malate sensitivity and phosphorylation state of PEPCase with temperature are of physiological significance. When the phosphorylated form of PEPCase is stabilized by reducing the temperature of leaves 9 h after transfer to constant darkness at 15 °C, a prolonged period of CO₂ fixation follows. When leaves are maintained in constant darkness at 15 °C until CO₂ output reaches a low steady-state level and the PEPCase is dephosphorylated, reducing the temperature to 3 °C results in a further period of CO₂ fixation even though the phosphorylation state of PEPCase does not change.Abbreviations CAM Crassulacean acid metabolism - PEP phosphoenolpyruvate - PEPCase phosphoenolpyruvate carboxylase We thank the Agricultural and Food Research Council for financial support for this work.     

Variation in coding exons of two electrophoretic alleles at the pigtail macaque carbonic anhydrase I locus as determined by direct,double-stranded sequencing of polymerase chain reaction (PCR) products

Two, electrophoretically distinct, forms of carbonic anhydrase I (CA Ia and CA Ib) are found at high polymorphic frequencies in red cells of natural populations of pigtail macaques,Macaca nemestrina, from southeast Asia. By use of the polymerase chain reaction, exons of the CA I gene were amplified from homozygous (a/a, b/b) and heterozygous (a/b) animals. Direct sequencing of the amplified DNA from four animals revealed differences between the a and the b electrophoretic alleles ranging from three to six nucleotides, and from one to three differences within each allele. These results indicate a greater genetic variability at the CA I locus in this macaque species than previously realized. We thank the personnel of the University of Washington, Regional Primate Research Center, Seattle, for sending us blood and tissue samples from pigtail macaques (supported by NIH Grant RR 00166). Our work was supported by NIH Grant GM 24681.     

Cytochemical and immunofluorescence investigations on insulin-like producing cells in the intestine of Mytilus edulis L. (Bivalvia)

Summary Insulin-like immunoreactivity can be localized to cells of the intestine in the-area of the hepatopancreas of Mytilus edulis L. No cross-reactivity can be obtained with anti-glucagon, anti-gastrin, anti-pentagastrin or anticaerulin. The cells containing the substance immunoreactive to mammalian anti-insulin, can be restained with paraldehyde-fuchsin.I thank Prof. A.G.E. Pearse for the opportunity to carry out this work in the Department of Histochemistry, Royal Postgraduate Medical School, London, England. The work was carried out with the aid of a grant from the Studienstiftung des Deutschen Volkes e.V., Germany (FRG).Dedicated to Prof. Dr. Drs. h.c. W. Bargmann at his 70th birthday.