bluuebunny/crossref_metadata_2025 huggingface dataset:
Imaging neuromuscular junctions by confocal fluorescence microscopy: individual endplates seen in whole muscles with vital intracellular staining of the nerve terminals
Authors: MARQUES M. J. , NETO H. SANTO
Published: March 2003

The mammalian neuromuscular junction has been extensively studied by different methods to understand better the biological aspects of its normal development, ageing and pathological conditions, such as disorders of neuromuscular transmission. In the present report, a new technique is described that combines confocal microscopy with the use of a vital nerve terminal dye (4‐Di‐2‐ASP) and rhodamine‐alpha‐bungarotoxin to stain postsynaptic acetylcholine receptors in the same endplate. Nerve terminals in the sternomastoid muscles of living adult mice were stained with 4‐Di‐2‐ASP, which labels intracellular compartments of the nerve terminal containing mitochondria. Slides of these muscles were viewed by confocal microscopy and images were stored on magnetic optical discs. This procedure was compatible with subsequent acetylcholine receptor staining with rhodamine‐α‐bungarotoxin and observation under the confocal microscope. Classical features of the adult neuromuscular junction were displayed, such as the branched‐pattern distribution of the nerve terminals and receptors and their complete colocalisation. In addition, nerve fibres from intramuscular nerve branches with their neighbouring cells, nuclei and muscle fibre striations could also be visualised. We conclude that the present technique can complement existing methods of investigation of nerve terminal anatomy and pathology, particularly where preservation of 3‐dimensional relationships is required and intracellular disturbances involving mitochondrial organisation, such as ageing or other degenerative disorders, may be present.
ARCHIBALD YOUNG 1913–1996
Authors: SHAW‐DUNN JOHN
Published: March 2003

Archibald Young was born in 1913 into a distinguished medical family. His father was a surgeon of international repute who followed Macewen to the Regius Chair of Surgery in Glasgow. On his mother's side he claimed descent from the famous Highland cateran Rob Roy. The family lived in great style in the fashionable Park area of Glasgow, and entertained Einstein on his visit to the University.

Young was educated at the High School of Glasgow and at St John's College Cambridge before returning to Glasgow to complete his medical training and take a number of junior clinical appointments in surgery and septics. When the Second World War broke out Young joined the RAMC and saw active service in North Africa and Italy. At the end of hostilities he was appointed to the Staff of the Anatomy Department in Glasgow, which remained his base for most of his career.
Immunohistochemical localisation of regulatory neuropeptides in human circumvallate papillae
Authors: KUSAKABE T. , MATSUDA H. , GONO Y. , FURUKAWA M. , HIRUMA H. , KAWAKAMI T. , TSUKUDA M. , TAKENAKA T.
Published: March 2003

The occurrence and distribution of neuropeptide‐containing nerve fibres in the human circumvallate papillae were examined by the peroxidase–antiperoxidase immunolocalisation method using surgical specimens that had not been subjected to radiotherapy, and the abundance of neuropeptide‐containing fibres was expressed as the percentage of total nerve fibres demonstrated by protein gene product (PGP) 9.5 immunoreactivity for a quantitative representation of these peptidergic fibres. Substance P (SP) and calcitonin gene‐related peptide (CGRP) immunoreactive (IR) nerve fibres were densely distributed in the connective tissue core of the circumvallate papillae, and some SP and CGRP‐IR fibres were associated with the taste buds. A moderate number of vasoactive intestinal polypeptide (VIP)‐IR fibres and a few galanin (GAL)‐IR fibres were also seen in the connective tissue core and subepithelial layer. There were, however, no VIP‐IR or GAL‐IR fibres associated with the taste buds. Neuropeptide Y (NPY)‐IR fibres were few and were associated with the blood vessels. Within the epithelium of the circumvallate papillae, no peptidergic fibres were found, although a number of PGP 9.5‐IR fibres were detected. The abundance of SP, CGRP, VIP, and GAL‐IR fibres expressed as the percentage of total PGP 9.5 IR fibres was 25.35±3.45%, 22.18±3.26%, 10.23±1.18%, and 4.12±1.05%, respectively. The percentage of NPY‐IR fibres was below 3%. In a deeper layer of the papillae, a few VIP, GAL, and NPY‐IR ganglion cells were found, and VIP immunoreactivity was detected in a few cells of the taste buds. There was no somatostatin, leucine enkephalin, or methionine enkephalin immunoreactivity in the circumvallate papillae. These results suggest that the dense SP and CGRP‐IR fibres within the connective tissue core of the human circumvallate papillae may be involved in the deep sensation of the tongue.
The possibility of deep peroneal nerve neurotisation by the superficial peroneal nerve: an anatomical approach
Authors: BÜYÜKMUMCU M. , ÜSTÜN M. E. , ŞEKER M. , KOCAOĞULLARI Y. , SAĞMANLIGİL A.
Published: March 2003

Neurotisation involves transfer of nerves for the restoration of function following injury. A number of nerves have been used in different part of the peripheral nervous system. This study was undertaken to develop a practical and relatively safe surgical approach to the treatment of L4 root lesion's. We examined the effectiveness and safety of neurotisation of the deep peroneal nerve and its branches by the superficial peroneal nerve. Twelve legs of dissected cadavers provided for teaching purposes in the anatomy laboratory were used to display the common peroneal nerve and its branches. Each branch was measured using calipers and analysed to investigate the possibility of neurotisation of the deep peroneal nerve by the superficial peroneal nerve and its branches. It was found that of the measured branches, transposition was possible between those to peroneus longus and tibialis anterior on the basis of their diameter and length. In recent decades, advances in microsurgical reconstruction and understanding of the microanatomy have played major roles in improving the results of surgical treatment of nerve injuries. There is a need for further experimental studies on the feasibility of this surgical approach.
Lectin binding patterns in nonsensory regions of rat cochlea during postnatal development
Authors: KHAN KHALID M. , SARFARAZ NOORJEHAN , SALIM ZEENAT
Published: March 2003

The distribution of glycoconjugates was examined in the nonsensory regions of the rat cochlea during postnatal development using biotin‐conjugated lectins. Temporal bones of rats at postnatal d 1 and at wk 2, 4 and 6 were fixed in 4% paraformaldehyde and 0.1% glutaraldehyde and processed for paraffin wax embedding. The dewaxed sections were incubated with 7 biotinylated lectins, followed by avidin‐biotin‐peroxidase complex. A different staining pattern was observed in the stria vascularis, spiral ligament and spiral limbus in the age groups examined. The staining intensity varied between lectins and the reaction product exhibited limited disparity. The staining intensity for WGA increased with age in all the 3 nonsensory regions. The staining patterns for the other lectins differed in the various nonsensory regions examined indicating tissue specificity. The limited variations in the lectin binding patterns after 2nd wk of postnatal life also indicate that the changes in the carbohydrate moieties are established during the fetal period of cochlear development and limited changes take place during postnatal maturation of the nonsensory regions.
Fibre composition in the interosseous nerve of the pigeon
Authors: NECKER REINHOLD , ROSENBERG JÖRG
Published: March 2003

The interosseous nerve of birds innervates a string of Herbst corpuscles located near the interosseous membrane between the tibia and fibula. Fibre composition of this nerve was assessed including both myelinated and unmyelinated axons. The diameter of the whole nerve is ∼100 μm. Complete data were obtained for 3 nerves. The mean total number of myelinated fibres and unmyelinated axons was 2872±53. The mean number of myelinated fibres was 280±20 and that for unmyelinated axons was 2600±47. There was a broad distribution of diameters for myelinated fibres ranging from ∼2 μm to 10 μm with a distinct peak at ∼3–5 μm and a less prominent second peak at 6–8 μm. Similarly, myelin sheath thickness distribution showed 2 peaks, one at 0.6–0.8 μm and another at 1.4–1.6 μm. It is suggested that the group represented by the second peak innervates the Herbst corpuscles. The group of smaller myelinated fibres and the unmyelinated axons are assumed to innervate other types of receptors, some of which may be nociceptors.
An unusual branching pattern of the superficial brachial artery accompanied by an ulnar nerve with two roots
Authors: ANAGNOSTOPOULOU S. , VENIERATOS D.
Published: March 2003

Variations in the arterial pattern of the upper limb are common and have been reported by several investigators (Fuss et al. 1985; Poteat, 1986; Tountas & Bergman, 1993; Rodriguez‐Baeza et al. 1995). These variations are often associated with anomalies in the arrangement of the nerves of the brachial plexus (Miller, 1939; Lengele & Dhem, 1989). The presence of a superficial brachial artery (Schwyzer & De Garis, 1935; Skopakoff, 1959; Fuss et al. 1985) and the usual pattern of its branching in the upper arm or forearm have also been reported (McCormack et al. 1953; Keen, 1961; Karlson & Niechalev, 1982; Lippert & Pabst, 1985; Rodriguez‐Baeza et al. 1995). The great variability of this arterial pattern may be attributed to the failure of regression of some paths of the embryonic arterial trunks (Tountas & Bergman, 1993; Rodriguez‐Baeza et al. 1995).

The aim of the present report is to describe the concomitant appearance of 3 unusual variations in the same upper limb of a male cadaver. In this arm: (1) a superficial brachial artery terminated its course by dividing into 3 branches at the cubital fossa; (2) the definitive brachial artery had an unusual origin; and (3) the ulnar nerve was abnormally formed from 2 roots. This novel variation is compared with other anatomical variations in the arterial supply of the upper limb. In a series of routine dissections of 100 embalmed human cadavers the following variations were observed in the right upper limb of a male subject.
Colocalisation of neuropeptides, nitric oxide synthase and immunomarkers for catecholamines in nerve fibres of the adult human vas deferens
Authors: JEN P. Y. P. , DIXON J. S. , GOSLING J. A.
Published: March 2003

Single and double‐label immunofluorescence methods were used to determine the distribution and patterns of colocalisation of various neuropeptides and nitric oxide synthase (NOS) with the catecholamine synthesising enzymes tyrosine hydroxylase (TH) and dopamine‐beta‐hydroxylase (DβH) in nerve fibres within specimens of adult human vas deferens obtained at vasectomy (age range 28 to 83 y). Cholinergic nerve fibres were immunolabelled with an antiserum to vesicular acetylcholine transporter (VAChT). Using the general nerve marker protein gene product 9.5 (PGP) the density of intramural nerve fibres was found to be similar irrespective of age. Many of these axons, especially in the outer 2 muscle layers were TH and DbH‐immunoreactive (IR) and were thus confirmed as noradrenergic. Fewer such axons were seen in the inner longitudinal muscle layer. All the noradrenergic nerve fibres also displayed NPY‐immunoreactivity with minor populations containing galanin (GAL) or somatostatin (SOM). Nerve fibres lacking TH and DbH‐IR were immunoreactive for VAChT and were sparsely distributed throughout the 2 outer muscle layers but more numerous in the inner muscle layer. Nerves lacking TH and DbH were immunoreactive for NPY and some also contained NOS, VIP or CGRP. These results have been compared with those obtained previously from specimens of human neonatal and infant vas deferens where, in contrast to the present results, NOS and VIP were shown to be colocalised with TH in many of the intramuscular nerve fibres. It thus appears that NOS and VIP cease their coexistence with TH in intramuscular nerve fibres of the human vas deferens between the pre‐ and postpubertal states. In addition to the intramuscular nerve fibres a VAChT‐IR subepithelial nerve plexus occurs in the vas deferens and may control the secretory activity of the lining epithelium. Most of these subepithelial nerve fibres were immunoreactive for NPY and many also contained VIP while minor populations were immunoreactive for NOS, GAL, SOM or SP although fibres containing CGRP were not observed. The neuropeptide content of the subepithelial nerve plexus was similar to that observed in the infant, except for an increased density of VIP‐IR nerves, which may reflect greater activity of the lining epithelial cells in the adult vas deferens.
Positional relationships between the masticatory muscles and their innervating nerves with special reference to the lateral pterygoid and the midmedial and discotemporal muscle bundles of temporalis
Authors: AKITA KEIICHI , SHIMOKAWA TAKASHI , SATO TATSUO
Published: March 2003

For an accurate assessment of jaw movement, it is crucial to understand the comprehensive formation of the masticatory muscles with special reference to the relationship to the disc of the temporomandibular joint. Detailed dissection was performed on 26 head halves of 14 Japanese cadavers in order to obtain precise anatomical information of the positional relationships between the masticatory muscles and the branches of the mandibular nerve. After complete removal of the bony elements, the midmedial muscle bundle in all specimens and the discotemporal muscle bundle in 6 specimens, derivatives of the temporalis, which insert into the disc were observed. On the anterior area of the articular capsule and the disc of the temporomandibular joint, the upper head of the lateral pterygoid, the midmedial muscle bundle of temporalis and the discotemporal bundle of temporalis were attached mediolaterally, and in 3 specimens the posterosuperior margin of the zygomaticomandibularis was attached to the anterolateral area of the disc. It is suggested that these muscles and muscle bundles contribute to various mandibular movements. Although various patterns of the positional relationships between the muscles and muscle bundles and the their innervating nerves are observed in the present study, relative positional relationships of the muscles and muscle bundles and of nerves of the mandibular nerve are consistent. A possible scheme of the developmental formation of the masticatory muscles based on the findings of the positional relationships between the muscles and the nerves is presented.
Growth promoting effects of human placental lactogen during early organogenesis: a link to insulin‐like growth factors
Authors: KAGAN KARABULUT AHMET , LAYFIELD ROBERT , PRATTEN MARGARET K.
Published: March 2003

Many maternally derived factors may be involved in the regulation of embryonic growth but the control mechanisms involved are poorly understood. Human placental lactogen (hPL) has been implicated in playing a role in the control of embryonic growth. Several investigators suggested that there may be a possible link between the effects of this hormone and insulin‐like growth factors (IGFs). In order to determine the growth promoting potential of hPL and involvement of IGFs in the mechanism of action of the hormone, 9.5 d rat embryos were cultured in vitro for 48 h in depleted serum in the presence and absence of hPL with additional IGF antisera. The growth supporting capacity of the serum was reduced by removal of low molecular weight molecules by prolonged filtration of the serum using filters with a molecular weight exclusion of 30 kDa. Addition of hPL (3.2–25.6 ng/ml) to depleted serum significantly improved embryonic growth and development, suggesting that the developing embryo may utilise hPL. The presence of antisera against hPL, IGF‐I and ‐II abolished the hPL‐induced increase in the development in all parameters suggesting that there may be a possible link between the IGFs and the effects of hPL on rat embryonic development and this hormone may achieve its growth promoting effects via IGFs.
Expression of transforming growth factor‐beta isoforms and their receptors in chronic tendinosis
Authors: FENWICK S. A. , CURRY V. , HARRALL R. L. , HAZLEMAN B. L. , HACKNEY R. , RILEY G. P.
Published: March 2003

Chronic tendon lesions are degenerative conditions and may represent a failure to repair or remodel the extracellular matrix after repeated micro‐injury. Since TGF‐β is strongly associated with tissue repair, we investigated the expression of TGF‐β isoforms (β1, β2 and β3) and their 2 signalling receptors (TGF‐βRI and TGF‐βRII) in normal and pathological Achilles tendons. In all tissues, all 3 TGF‐β isoforms and the 2 receptors were present at sites of blood vessels. Cells in the matrix showed no staining for TGF‐β1 or β3, while TGF‐β2 was associated with cells throughout the normal cadaver tendon. Tissue from tendons with pathological lesions showed an increase in cell numbers and percentage TGF‐β2 expression. TGF‐βRII showed a wide distribution in cells throughout the tissue sections. As with TGF‐β2, there was an increase in the number of cells expressing TGF‐βRII in pathological tissue. TGF‐βRI was restricted to blood vessels and was absent from the fibrillar matrix. We conclude that despite the presence and upregulation of TGF‐β2, TGF‐β signalling is not propagated due to the lack of TGF‐βRI. This might explain why chronic tendon lesions fail to resolve and suggests that the addition of exogenous TGF‐β will have little effect on chronic tendinopathy.
Demonstration of Birbeck (Langerhans cells) granules in the normal chicken epidermis
Authors: PÉREZ‐TORRES ARMANDO , USTARROZ‐CANO MARTHA
Published: March 2003

Mammalian Langerhans cells (LC) are epidermal dendritic cells which originate in bone marrow and migrate toward the T cell area of lymph nodes, where they act as professional antigen‐presenting cells. A variety of cell surface markers, such as the ectoenzyme adenosine triphosphatase (ATPase), Ia and CD1a antigens, have been used extensively to identify LC. Ultrastructural identification of this cell type in the mammalian epidermis is made by the demonstration of a typical and unique cytoplasmic organelle, the Birbeck granule (BG). Although we had earlier demonstrated the coexpression of ATPase and Ia antigens on epidermal dendritic cells of the chicken epidermis, the presence of the BG has not previously been documented. The aim of the present study was to investigate whether chicken epidermal LC‐like cells possess an organelle similar to the BG, and thus to complete their identification. Our findings are the first demonstration of characteristic rod‐shaped, racket‐shaped and disc‐shaped intracytoplasmic organelles, morphologically similar to the mammalian BG, in avian LC.
Regulation of microvascular permeability by vascular endothelial growth factors*
Authors: Bates D. O. , Hillman N. J. , Williams B. , Neal C. R. , Pocock T. M.
Published: March 2003

Generation of new blood vessels from pre‐existing vasculature (angiogenesis) is accompanied in almost all states by increased vascular permeability. This is true in physiological as well as pathological angiogenesis, but is more marked during disease states. Physiological angiogenesis occurs during tissue growth and repair in adult tissues, as well as during development. Pathological angiogenesis is seen in a wide variety of diseases, which include all the major causes of mortality in the west: heart disease, cancer, stroke, vascular disease and diabetes. Angiogenesis is regulated by vascular growth factors, particularly the vascular endothelial growth factor family of proteins (VEGF). These act on two specific receptors in the vascular system (VEGF‐R1 and 2) to stimulate new vessel growth. VEGFs also directly stimulate increased vascular permeability to water and large‐molecular‐weight proteins. We have shown that VEGFs increase vascular permeability in mesenteric microvessels by stimulation of tyrosine autophosphorylation of VEGF‐R2 on endothelial cells, and subsequent activation of phospholipase C (PLC). This in turn causes increased production of diacylglycerol (DAG) that results in influx of calcium across the plasma membrane through store‐independent cation channels. We have proposed that this influx is through DAG‐mediated TRP channels. It is not known how this results in increased vascular permeability in endothelial cells in vivo. It has been shown, however, that VEGF can stimulate formation of a variety of pathways through the endothelial cell, including transcellular gaps, vesiculovacuolar organelle formation, and fenestrations. A hypothesis is outlined that suggests that these all may be part of the same process.
Constitutive expression of the low‐affinity neurotrophin receptor and changes during axotomy‐induced death of sensory neurones in the neonatal rat dorsal root ganglion
Authors: Murray Simon S. , Cheema Surindar S.
Published: March 2003

Sensory neurones in the dorsal root ganglion (DRG) of the neonatal rat express the 75‐kDa low‐affinity neurotrophin receptor (p75NTR) and these neurones degenerate rapidly after axotomy. p75NTR belongs to the tumour necrosis factor superfamily, several members of which have a role in cell death and it is constitutively expressed within a subpopulation of DRG neurones. p75NTR has been implicated in mediating the degeneration of these neurones after axotomy. In this study, we characterize the expression of p75NTR in sensory neurones of the newborn rat DRG using immunohistochemistry. Furthermore, we investigate the change in constitutive expression pattern of p75NTR in these neurones following axotomy. In the C7 and C8 DRG of the newborn rat, p75NTR is expressed in approximately 70% of DRG neurones. Those expressing p75NTR can be classified into subpopulations with moderate or intense p75NTR expression, each present in approximately equal proportions. Whilst p75NTR expression is observed in neurones throughout the entire neuronal diameter range, a correlation exists between neuronal diameter and p75NTR expression intensity. We also found that the most vulnerable population following axotomy were those sensory neurones which constitutively express the highest levels of p75NTR, i.e. the large‐diameter neurones.
The response of perennial ryegrass/white clover swards to elevated atmospheric CO2concentrations 1. Effects on competition and species composition and interaction with N supply
Authors: SCHENK U. , JÄGER H.‐J. , WEIGEL H.‐J.
Published: March 2003

The effects of long‐term carbon dioxide enrichment on competition for nutrients and light in a ryegrass/clover association were determined for simulated swards of perennial ryegrass (Lolium perenne L. cv. Parcour) and white clover (Trifolium repens L. cv. Karina), which were grown as monocultures and in three mixtures (25/75, 50/50, 75/25), according to the replacement design, at two levels of nitrogen (N) supply (no additional N and 200 kg N ha−1) and at season‐long ambient (380 ppm) and elevated (670 ppm) CO2 concentrations, in open‐top chambers. Stands were cut four times, at about monthly intervals, to a height of 5 cm. Plant material was separated into different species, fresh and dry weights were determined and the content of macroelements (N, P, K, S, Mg) in both species was measured. In addition, plant height of both species at harvest dates and during several regrowth periods was monitored.

Results indicate that both species made demand on different resources and profited from growth in a mixed sward. Co2 related yield increase amounted to 16–4–2 % for white clover whereas the effect of high CO2 on ryegrass yield ranged between −33% and +9% depending on N supply, mixture and year. As a result the contribution of white clover to total yield in mixed swards was significantly enhanced by CO2 enrichment at many harvests in both N supply treatments. Without additional N supply, shoot competition for light was intensified by CO2 enrichment to the disadvantage of ryegrass, since clover petioles grew longer and ryegrass was shorter at elevated CO2 With N fertilization, no marked effect of CO2 enrichment on interspecific competition could be observed. Since clover and total yield were increased by CO2 enrichment, nutrient requirements were also increased and potassium deficiency and increased intraspecific competition of clover for K was observed in the mixtures under elevated CO2 which had the highest nutrient withdrawal. Although white clover profited much more from CO2 enrichment in both N fertilization treatments, the suppression of ryegrass in mixed swards could only be observed under low N conditions.

Generally, the effect of N fertilization on competitive interference between both species was much greater than the effect of CO2 enrichment and it is suggested that the effect of elevated CO2 on the balance of species and the outcome of competition in a grass/clover sward is mainly dependent on the N status.
Evidence that ozone exposure increases the susceptibility of plants to natural frosting episodes
Authors: FOOT J. P. , CAPORN S. J. M. , LEE J. A. , ASHENDEN T. W.
Published: March 2003

This study investigates the phytotoxicity of ozone during periods of natural frosts following long‐term fumigation. Heather (Calluna vulgaris (L.) Hull.) plants were fumigated with ozone for 6 months at 70 nl I−1, for 8 h d−1, 5 d a week in open‐top chambers. Natural frosting occurred within the chambers towards the end of this period (5–27 December 1995). Plant responses to the ozone/natural frosting episode were recorded in January 1996. We show conclusively that naturally occurring frost exposure results in increased damage and mortality for ozone‐treated plants, which could have important implications for the function of heathland ecosystems.
A comparison of the vegetative growth of male‐sterile and hermaphroditic lines of Plantago lanceolata in relation to N supply
Authors: POOT PIETER , VAN DEN BROEK TOMMY , VAN DAMME JOS M. M. , LAMBERS HANS
Published: March 2003

summary

Gynodioecy is a dimorphic breeding system in which hermaphrodites coexist with male sterlies in natural populations. Theoretical models predict that without any compensation in female fitness, male sterlies will quickly disappear from a population. The amount of compensation required depends upon the mode of inheritance. In this study we investigated whether performance, during early vegetative growth, could play a role in the maintenance of male sterlies in populations of gynodioecious self‐incompatible Plantago lanceolata. This was accomplished by comparing the growth of the predominant male‐sterile type from a natural population with two hermaphroditic types. One of the hermaphroditic types differed from the male sterile only in nuclear genome, having the same type of cytoplasm. The other type was nuclearly nearly isogenic, but had a different cytoplasm. Plants were grown under controlled conditions, either in near‐optimal hydroponic solutions, or in a range of N‐supplies on sand. A detailed growth analysis was carried out, and the relative growth rate (RGR) of each type was analysed into its underlying components, the net assimilation rate, the leaf mass ratio and the specific leaf area. No difference between the sex types in relative growth rate was found and, in the components underlying the RGR, only a few small differences were detected. In none of the growth parameters examined did the male sterlies differ from either hermaphroditic type. Differences in plant growth related to N‐supply, differences in growth on hydroponics compared with sand culture, as well as the maintenance of male sterility in populations of P. lanceolata are discussed.
Cytosolic expression of the Bacillus amyloliquefaciens SacB protein inhibits tissue development in transgenic tobacco and potato
Authors: CAIMI P. G. , McCOLE L. M. , KLEIN T. M. , HERSHEY H. P.
Published: March 2003

Fructans are linear or branched polymers containing a single sucrose and repeating fructose residues. An early model for fructan biosynthesis in higher plants suggested that partial synthesis of the polymer occurred in the cell cytosol. The current model suggests that synthesis requires the interaction of two separate fructosyltransferases located in the vacuole. Tobacco lines containing a chemically induced promoter, directing expression of the Bacillus amyloliquefaciens SacB gene in the present study, provided an opportunity to regulate and target fructan synthesis to the cytosol of transgenic plants. Induced expression of the gene led to rapid destruction of leaf tissue. Amino acid substitution at a highly conserved site (Arg331) in the SacB gene reduced the fructosyltransferase efficiency without reducing the invertase activity of the enzyme. Expression of the mutant gene in transgenic tobacco also resulted in leaf damage. However, the appearance of necrotic tissue was greatly delayed. The results suggest that the phenotype is due to accumulation of fructan in the cytosol. Fructan metabolism in the cytosol of potato tubers was also detrimental to development. Tuber size and starch synthesis was significantly reduced in lines containing the untargeted gene. Transgenic tobacco and potato containing the SacB gene offer an opportunity to study the metabolism of fructan and the effect of accumulation on plant cell development.
Synthesis of fructans by partially purified fructosyltransferase activities from Lolium rigidum
Authors: ST. JOHN J. A. , SIMS I. M. , BONNETT G. D. , SIMPSON R. J.
Published: March 2003

Sucrose∶sucrose fructosyltransferase (SST) activity was partially purified from whole shoots of Lolium rigidum by a combination of affinity chromatography, gel filtration and anion‐exchange chromatography. The SST activity co‐eluted with some fructan∶fructan fructosyltransferase (FFT) and invertase activities and consequently the partially purified preparation was termed the fructosyltransferase (FT) preparation.

The SST‐like activity in the FT preparation was purified 214‐fold and had an apparent molecular mass of 84000. The FT preparation contained several peptides with an apparent pI of 4·6–4·7. When assayed with sucrose concentrations up to 600 mM, the FT preparation synthesized 1‐kestose at all concentrations, and synthesized 6‐kestose at concentrations of 150 mM and greater. The Km of 1‐kestose production was 0·2 M. When the FT preparation was assayed at a concentration of activity approximately half that measured in fresh tissue with 100 mM sucrose, 1‐kestose, or 6G‐kestose as substrates, fructans of degree of polymerization (DP) 5 were synthesized.

A partially purified FFT activity, free of SST and invertase activities, which synthesized β‐2,1‐glycosidic linked oligofructans of DP 6, was combined in vitro with the FT preparation (FFT‐FT preparation) to give a ratio of SST∶FFT activities similar to that measured in crude enzyme extracts from L. rigidum. The FFT‐FT preparation synthesized oligofructans when assayed with 100 mM concentrations of sucrose, 1‐kestose or 6G‐kestose, but was not able to synthesize fructans of DP 6 even after extended assays of up to 10 h. The FFT‐FT preparation was also assayed with 100 mM sucrose with small amounts of concentrated sucrose added periodically during the assay to maintain the substrate concentration. In this assay, the FFT‐FT preparation synthesized fructans up to an apparent DP of 17 or greater. The fructans of DP 6 synthesized in the assay appeared to form two molecular series containing both β‐2,1‐ and β‐2,6‐glycosidic linked fructosyl residues with terminal or internal glucosyl residues. The apparent rate of SST activity in the assay of the FFT‐FT preparation was greater than that measured in a similar assay of the FT preparation alone which did not result in fructans with DP 6. It was concluded that the FFT‐FT preparation, when assayed with a continual supply of sucrose, contained a factor which promoted synthesis of fructans of DP 6 and synthesis of β‐2,6‐glycosidic linkages between fructosyl residues.
Thioredoxins: structure and function in plant cells
Authors: JACQUOT JEAN‐PIERRE , LANCELIN JEAN‐MARC , MEYER YVES
Published: March 2003

Thioredoxins are ubiquitous small‐molecular‐weight proteins (typically 100–120 amino‐acid residues) containing an extremely reactive disulphide bridge with a highly conserved sequence ‐Cys‐Gly(Ala/Pro)‐Pro‐Cys‐. In bacteria and animal cells, thioredoxins participate in multiple reactions which require reduction of disulphide bonds on selected target proteins/ enzymes. There is now ample biochemical evidence that thioredoxins exert very specific functions in plants, the best documented being the redox regulation of chloroplast enzymes. Another area in which thioredoxins are believed to play a prominent role is in reserve protein mobilization during the process of germination. It has been discovered that thioredoxins constitute a large multigene family in plants with different‐subcellular localizations, a unique feature in living cells so far. Evolutionary studies based on these molecules will be discussed, as well as the available biochemical and genetic evidence related to their functions in plant cells. Eukaryotic photosynthetic plant cells are also unique in that they possess two different reducing systems, one extrachloroplastic dependent on NADPH as an electron donor, and the other one chloroplastic, dependent on photoreduced ferredoxin. This review will examine in detail the latest progresses in the area of thioredoxin structural biology in plants, this protein being an excellent model for this purpose. The structural features of the reducing enzymes ferredoxin thioredoxin reductase and NADPH thioredoxin reductase will also be described. The properties of the target enzymes known so far in plants will be detailed with special emphasis on the structural features which make them redox regulatory. Based on sequence analysis, evidence will be presented that redox regulation of enzymes of the biosynthetic pathways first appeared in cyanobacteria possibly as a way to cope with the oxidants produced by oxygenic photosynthesis. It became more elaborate in the chloroplasts of higher plants where a co‐ordinated functioning of the chloroplastic and extra chloroplastic metabolisms is required.

CONTENTSSummary543I.Introduction544II.Thioredoxins from photosynthetic organisms as a structural model545III.Physiological functions552IV.The thioredoxin reduction systems556V.Structural aspects of target enzymes558VI.Concluding remarks563Acknowledgements564References564
Effect of aluminium on ion uptake and H+ release by maize
Authors: CALBA HENRI , JAILLARD BENOÎT
Published: March 2003

summary

The aim of this study was to quantify the long‐term effects of Al in solution on mineral ion uptake and associated H+ release by the roots of maize cultivated in conditions compatible with field data. Two maize (Zea mays L.) cvs of differing sensitivity to Al were compared. The culture was conducted in hydroponic conditions for culture periods of 2–5 d in the presence of 0–30 μM Al. Proton fluxes were localized along the roots using a videodensitometry method with bromocresol green on agarose gel.

Aluminium reduced the uptake of anions, particularly NO3−, whereas the uptake of K+ and NH4+ was unaffected. These effects were accompanied by an increase in net H+ release which was quantitatively comparable with the reduction in NO2− uptake. The reduction in NO3− uptake and the concomitant increase in H+ release increased with the quantity of Al accumulated in the roots, which was itself dependent on the concentration of Al in solution. The Al‐sensitive cultivar bound more Al to its roots than the Al‐tolerant one; the reduction in NO3− uptake and the increase in H+ release were also greater in the Al‐sensitive cv. Dye indicator videodensitometry showed that the increased H+ release was general all along the roots.

These results led us to specify the relation between the Al deposition in roots, the reduction in NO3− uptake and the stimulation in rhizosphere acidification by the plants. They show that, with maize, Al induces an inhibition in NO3− uptake and an increase in net H+ release in a 1:1 ratio. These concomitant effects suggest that inhibition in NO3−/OH− exchange unmasks active H+ excretion which is otherwise insensitive to Al. Both these effects are closely related to the Al content in roots expressed per unit root length.
Oxides of nitrogen and ozone: can our plants survive?
Authors: LEA PETER J.
Published: March 2003

The authoritative talk by Professor Fowler (Fowler et al., 1998), emphasized the huge increase in the rate of NOx (NO and NO2) emissions into the atmosphere due to fossil fuel combustion, from 1 Tg N y−1 to over 20 Tg N y−1 during the 100 yr between 1880 and 1980. He went on to predict that this rate of emission from anthropogenic sources would increase to 46 Tg N y−1 by the year 2025. In addition, NO can also be released from the soil following microbial action, a process that is very dependent upon soil temperature, nitrogen availability and water content. Later in the meeting, Professor Raven (Raven & Yin, 1998) pointed out that terrestrial plants, though not necessarily each individual species, have over the past 450 million yr coped with large changes in nitrogenous compounds in the environment. Nevertheless, this is no basis for complacency about the current situation because the rates of change caused by man's activities are probably unprecedented. Furthermore, the fact that terrestrial plant life in some form can continue, despite massive changes in environmental chemistry, does not necessarily indicate that the systems on which we ourselves are dependent will be conserved.
Uptake and fate of gaseous pollutants in leaves
Authors: CAPE J. N.
Published: March 2003

The final paper of the Symposium (delivered by Professor Raven) reviewed the rôle of nitrogen in the development of terrestrial life, and gave a timely reminder of the context of the current debate on the effects of anthropogenic N; current man‐made perturbations of the N cycle are approaching the scale of catastrophic perturbations caused by major meteorite impacts.

Faced with such a stark evaluation of the current position one is forced to consider how the scientific community is reacting to the challenge of understanding and predicting the effects of man's activities.
Nitrogen on microbial and global scales
Authors: VAN DER EERDEN LUDGER
Published: March 2003

Several temporal and spatial scales have been considered during the Symposium, and the presentations by Wallenda & Kottke (1998) and by Norby (1998) represent two extreme points in a wide range. They both deal with short and long‐term impacts on the local as well as the global scale. I present here a few additional thoughts, and a subject that both papers have touched only slightly will be expanded: the differential impact of different forms of nitrogen deposition.
Atmospheric ammonia and impacts of nitrogen deposition: uncertainties and challenges
Authors: SCHJØRRING JAN K.
Published: March 2003

Dr Willem Asman concluded that the major global sources of atmospheric NH3 are excreta from domestic animals and fertilizers. A question raised was: how reliable are the emission estimates and extrapolations? The answer was that emission estimates are surrounded by uncertainty, which is a major handicap to sound modelling of NH3 dry deposition and, consequently, to obtaining good estimates of critical load exceedences.

Major uncertainties in emission estimates seem to be related to the use of simple emission factors, many of which are highly empirical or have been derived from measurements carried out under conditions which deviate considerably from those following modern practices of handling and applying manure and fertilizers. An example is provided by the commonly used emission factors for synthetic fertilizers (see e.g. Bouwman et al. (1997)), which are much higher than recent micrometeorological assessments seem to suggest. Thus, emission from urea, the most widespread fertilizer used in the world (currently around 55% of world N consumption) can be completely avoided if the fertilizer is incorporated into the upper soil layers. Similarly, a growing crop can reduce losses to well below 10% of the applied amount of urea‐N, i.e. to less than half of the generally used emission factors of 15% for Europe and 25% for the tropics. The emission factor for NPK‐fertilizer is set at 4%, whereas that for pure calcium‐ammonium‐nitrate, the same N compound as is present in NPK‐fertilizers, is assumed to be only 2%.
Nitrogen deposition and ectomycorrhizas
Authors: WALLENDA THOMAS , KOTTKE INGRID
Published: March 2003

As a result of increasing anthropogenic nitrogen deposition, N availability in many forest ecosystems, which are normally N‐limited, has been enhanced. We discuss the impacts of this increased N availability on the ectomycorrhizal (ECM) symbiosis which is generally regarded as an adaptation to nutrient limited conditions. Nitrogen deposition can influence fruit‐body formation by ECM fungi, the production and distribution of the extraradical mycelium in the soil and the formation of ectomycorrhizas.

Available data from long‐term N deposition studies indicate that the most prominent effects might be discernible above‐ground (i.e. on the formation of fruit bodies). ‘Generalist’ species, forming a symbiosis with a wide range of tree species, seem to be less affected by increased N availability than ‘specialist’ species, especially those living in symbiosis with conifers. However, the importance of below‐ground investigations to determine the impacts of N deposition on the ECM symbiosis must not be underestimated. Culture experiments show an optimum N concentration for the formation of extraradical mycelium and mycorrhizas. Often, negative effects only become visible at comparatively high N concentrations, but the use of a few easily cultivated species of ECM fungi, which are adapted to higher N concentrations, undermines our ability to generalize.

So far, N deposition experiments in the field have only shown minor changes in the below‐ground mycorrhizal population, as estimated from the investigation of mycorrhizal root tips. However, effects on the ECM mycelium, which is the main fungal component in terms of nutrient uptake, cannot be excluded and need further consideration.

Because the photoassimilate supply from the plant to the fungal partner is crucial for the maintenance of the ECM symbiosis, we discuss the possible physiological implications of increasing N inputs on the allocation of C to the fungus. Together with ultrastructural changes, physiological effects might precede obvious visible changes and might therefore be useful early indicators of negative impacts of increasing N inputs on the ECM symbiosis.
Nod factors and tri‐iodobenzoic acid stimulate mycorrhizal colonization and affect carbohydrate partitioning in mycorrhizal roots of Lablab purpureus
Authors: XIE ZHI‐PING , MÜLLER JOACHIM , WIEMKEN ANDRES , BROUGHTON WILLIAM J. , BOLLER THOMAS
Published: March 2003

Roots of Lablab purpureus (L.) Sweet were treated with tri‐iodobenzoic acid (TIBA), kinetin or with nodulation factors (Nod factors) purified from Rhizobium sp. NGR234 and grown in the presence of a mycorrhizal inoculum (Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe. Colonization by the mycorrhizal fungus was increased from <30% to c. 65% of root length when roots were treated with these growth regulators. Moreover, treatment of mycorrhizal L. purpureus roots with Nod factors or TIBA strongly induced sporocarp formation of Glomus mosseae. In parallel, the pool size of the fungal disaccharide trehalose was significantly affected in roots treated with TIBA and Nod factors alone, and with TIBA combined with all effectors, and increased from 0·06 mg g−1 d. wt in control roots to up to 1·7 mg g−1 d. wt (TIBA+kinetin). Conversely, the sucrose pool decreased from 5% d. wt to less than a half in roots treated with Nod factors. Activities of trehalase were significantly enhanced in mycorrhizal roots by the treatment with Nod factors or TIBA. When Nod factors and TIBA were added in combination, these activities were strongly enhanced suggesting synergism between these growth regulators.
Nitrate and phosphate uptake by Agropyron desertorum and Artemisia tridentata from soil patches with balanced and unbalanced nitrate and phosphate supply
Authors: CUI MUYI , CALDWELL MARTYN M.
Published: March 2003

To test the ability of plants to integrate small‐scale imbalances in soil nitrate and phosphate patches, plant growth and acquisition of nitrate and phosphate were measured for the perennial grass Agropyron desertorum (Fisch. ex Link) Schult. and the shrub Artemisia tridentata Nutt. ssp. vaseyana (Rydb.) Beetle in soil where the principal supply of nitrate and phosphate came from two enriched patches. The soil was calcareous loamy‐skeletal Typic Haploxerolls. These patches were applied in two treatments: either nitrate and phosphate were applied in both patches (balanced treatment) or one patch contained only nitrate and the other only phosphate (unbalanced treatment). The same total quantity of nutrients was applied in both treatments and these included 15N and 32P tracers. The plants were in large pots in open field conditions. There were no significant differences in total biomass production and nitrogen concentration between the two treatments, indicating that both species had the physiological ability to integrate soil nutrient resources. Artemisia was able to acquire more phosphate in the unbalanced treatment, probably due to the high local solution phosphate concentration. Generally Artemisia acquired more N and P than did Agropyron.
The role of mycorrhiza in determining the response of Plantago lanceolata to CO2 enrichment
Authors: ROUHIER HERVÉ , READ DAVID J.
Published: March 2003

Plantago lanceolata L. was grown for 104 d with (M) or without (NM) arbuscular mycorrhizal colonization under conditions of ambient (CAMB=350 μl l−1) and elevated (CELEV=540 μl l−1) CO2. Sequential harvests (H) were taken at 41 (H1), 76 (H2) and 104 d (H3) to determine the time‐course of mycorrhizal influence on the response of the plant to CO2 enrichment. Total yields of M plants were greater than those of NM from H2 onwards. Plants in the MELEV treatment were significantly larger than those in the MAMB at 104 d. There were significant but much smaller differences in yield between NMELEV and NMAMB. The differences in total yield arose through impact of CELEV on both shoots and roots. Total root length was greater in MELEV than in MAMB only at H3, but total length of mycorrhizal root was greater at H2 and H3. The percentages of root length colonized and that occupied by arbuscules and vesicles were greater in MELEV than in MAMB at the last two harvests, indicating increased sequestration of carbon in internal fungal structures. Though extraradical hyphal lengths were greater in MELEV than in MAMB at H2 and H3, the differences were not significant. Phosphorus inflow and P content of M plants were higher than those of NM plants at H2 and H3, and were higher in MELEV than in MAMB at H3. anova revealed no significant interactions between CO2 and mycorrhizal treatment.

The results are discussed in relation to carbon sequestration in mycorrhizal systems and likely impacts of CO2 enrichment on P. lanceolata grown under field conditions. The importance of sequential harvesting for realistic determination of responses to CO2 is stressed.