bluuebunny/crossref_metadata_2025 huggingface dataset:
Stomatal density and needle anatomy of Scots pine (Pinus sylvestris) are affected by elevated CO2
Authors: Lin Jinxing , Jach M. E. , Ceulemans R.
Published: March 2003

Morphological and anatomical characteristics of needles from different whorl levels were examined on Scots pines (Pinus sylvestris), grown for 4 yr under either ambient or elevated (ambient + 400 µmol mol−1) CO2 concentrations in open‐top chambers.

 Needle characteristics were studied using light microscopy, scanning electron microscopy and laser scanning confocal microscopy.

 Under the elevated CO2 treatment stomatal density was reduced on both adaxial and abaxial needle surfaces although the number of rows of stomata did not change significantly. Needle cross‐sectional area increased by 10%; this was largely the result of an increase in needle thickness and, to a lesser extent, needle width. The increase in needle thickness was due to a large increase in mesophyll tissue. The relative area (i.e. proportion of the total area) of epidermis plus hypodermis, of resin canal, of xylem and of central cylinder decreased, whereas the relative area of needle phloem significantly increased.

 The results suggest that a prolonged exposure to elevated CO2 has an effect on needle structure, anatomical and stomatal characteristics of Scots pine needles.
The CDPK superfamily of protein kinases
Authors: Harmon Alice C. , Gribskov Michael , Gubrium Erika , Harper Jeffrey F.
Published: March 2003

The CDPK superfamily consists of six types of protein kinases, which differ in the regulatory domains they contain. CDPKs (calcium‐dependent protein kinases or calmodulin‐like domain protein kinases) are activated by the binding of calcium to their calmodulin‐like regulatory domains. The carboxyl terminal domains of CRKs (CDPK‐related kinases) have sequence similarity to the regulatory domains of CDPKs, but do not bind calcium. PPCKs (PEP carboxylase kinases) contain only a catalytic domain. PRKs (PPCK‐related kinases) have a carboxyl‐terminal domain that has no similarity to that of any other member of the superfamily. CCaMKs (calcium and calmodulin regulated kinases) bind both calcium ions and the calcium/calmodulin complex, whereas CaMKs (calmodulin‐dependent protein kinases) bind the calcium/calmodulin complex, but not calcium. Phylogenetic trees constructed from amino acid sequences of catalytic or regulatory domains show that CDPKs and CRKs are closely related and might share a common ancestor. Plant CCaMKs and CaMK form a group more closely related to protozoan, than to plant, CDPKs. Intron analysis of the 42 CDPK, CRK, PPCK, and PRK genes from Arabidopsis supports the structure of the gene trees, the possibility that PPCKs/PRKs belong to the CDPK superfamily, and suggests that several introns have been added during evolution of the family.
Arsenic uptake and metabolism in arsenic resistant and nonresistant plant species
Authors: Meharg Andrew A. , Hartley‐Whitaker Jeanette
Published: March 2003

Elevation of arsenic levels in soils causes considerable concern with respect to plant uptake and subsequent entry into wildlife and human food chains. Arsenic speciation in the environment is complex, existing in both inorganic and organic forms, with interconversion between species regulated by biotic and abiotic processes. To understand and manage the risks posed by soil arsenic it is essential to know how arsenic is taken up by the roots and metabolized within plants. Some plant species exhibit phenotypic variation in response to arsenic species, which helps us to understand the toxicity of arsenic and the way in which plants have evolved arsenic resistances. This knowledge, for example, could be used produce plant cultivars that are more arsenic resistant or that have reduced arsenic uptake. This review synthesizes current knowledge on arsenic uptake, metabolism and toxicity for arsenic resistant and nonresistant plants, including the recently discovered phenomenon of arsenic hyperaccumulation in certain fern species. The reasons why plants accumulate and metabolize arsenic are considered in an evolutionary context.

Contents

Summary
29

I.
Introduction
30

II.
Toxicity of arsenic species to plants
33

III.
Arsenic resistance in plants
35

1. Arsenic resistance mechanisms
35

2. Arsenate resistence polymorphism
36

3. Mycorrhizal fungi and arsenic resistance
36

IV.
Complexation of arsenic
37

V.
Arsenic metabolism
37

VI.
Arsenic hyperaccumulation
39

VII.
Evolutionary context
40

VIII.
Conclusions
40

References
40
Is differential phytochelatin production related to decreased arsenate influx in arsenate tolerant Holcus lanatus?
Authors: Hartley‐Whitaker J. , Woods C. , Meharg A. A.
Published: March 2003

Phytochelatins (PCs) are required for arsenic (As) detoxification in nontolerant plants. In addition, a role for PCs in arsenate tolerance has recently been proven, with tolerant plants able to accumulate significantly higher concentrations of As–PC complexes at equivalent levels of stress than nontolerant plants.

The relationship between arsenate influx and PC production in tolerant and nontolerant Holcus lanatus plants was determined in this study, along with an investigation of the effect of inhibition of PC synthesis by buthionine sulfoximine (BSO) on arsenate tolerance.

A strong correlation between PC production and arsenate influx was demonstrated in arsenate‐tolerant plants. In addition, inhibition of PC synthesis by BSO in tolerant plants increased arsenate sensitivity to that of the nontolerant clone. This dramatic reduction in tolerance proves that PC production is an essential component of the arsenate tolerance mechanism in H. lanatus .

This study proposes that while there is a single major gene for arsenate tolerance, hypostatic modifiers are also in operation, affecting the expression of the tolerance character.
Carbon isotope ratio and the extent of daily CAM use by Bromeliaceae
Authors: Pierce Simon , Winter Klaus , Griffiths Howard
Published: March 2003

Use of carbon isotope ratio (δ 13
C) to resolve photosynthetic pathways (C
3
, C
4 or CAM) has limitations imposed by the use of intermediate photosynthetic modes by certain plant taxa.

Diel gas‐exchange patterns, leaf δ 13 C values and nocturnal tissue acidification were determined for 50 Bromeliaceae.

δ 13
C values for well watered plants reflected the proportion of daily CO
2
uptake occurring at night. Thirteen per cent of species with δ
13
C values typical of C
3
plants (i.e. from −22.6 to −31.5‰) showed nocturnal acidification and either a small proportion (< 10%) of daily CO
2
uptake occurring nocturnally or internal CO
2
recycling during part of the night. None altered CAM expression in response to short‐term drought, but the contribution of CAM to daily carbon gain became proportionally more important as C
3
CO
2 uptake failed.

Surveys of plant communities using solely the carbon isotope technique underestimate the number of CAM‐equipped plants.
Reactions of paratracheal cells of resistant and susceptible carnation (Dianthus caryophyllus) cultivars to vascular invasion by Fusarium oxysporum f. sp. dianthi
Authors: Ouellette G. B. , Baayen R. P. , Simard M. , Rioux D.
Published: March 2003

Here, cell and wall modifications in resistant (cv. I
) and susceptible (cv.
II
) carnation inoculated with
Fusarium oxysporum are characterized.

Samples, of various days after inoculation (dai), were double‐fixed and gold‐complexed probes used for cellulose, pectin, chitin.

In cv. I
, mature paratracheal cells contained appositions mostly bound by a cellulose layer over the densely opaque native walls. Middle lamellae of younger cells, many hyperplastic and much distorted, were obliterated, and appositions displayed a mingling of cellulose‐labelled and ‐unlabelled components; adjoining cells contained inner suberin layers. Cell wall modifications in cv.
II
were limited, but early pathogen invasion was restricted to middle lamellae despite pronounced host cell content alterations. In both cvs, microfilaments linked to often small, thin‐walled fungal cells in vessels and in turn to coating on their walls, extended into adjoining cells, seemingly associated in cv.
I
with cell reactions, and in cv.
II excreted as bodies into vacuoles.

The present results highlight that extensive cell reactions and wall modifications, including suberin deposition in hyperplastic tissue, are the main initial factors of defence conducive to final pathogen confinement.
Plant resistance towards insect herbivores: a dynamic interaction
Authors: Gatehouse John A.
Published: March 2003

Plant defences against insect herbivores can be divided into ‘static’ or constitutive defences, and ‘active’ or induced defences, although the insecticidal compounds or proteins involved are often the same. Induced defences have aspects common to all plants, whereas the accumulation of constitutive defences is species‐specific. Insect herbivores activate induced defences both locally and systemically by signalling pathways involving systemin, jasmonate, oligogalacturonic acid and hydrogen peroxide. Plants also respond to insect attack by producing volatiles, which can be used to deter herbivores, to communicate between parts of the plant, or between plants, to induce defence responses. Plant volatiles are also an important component in indirect defence. Herbivorous insects have adapted to tolerate plant defences, and such adaptations can also be constitutive or induced. Insects whose plant host range is limited are more likely to show constitutive adaptation to the insecticidal compounds they will encounter, whereas insects which feed on a wide range of plant species often use induced adaptations to overcome plant defences. Both plant defence and insect adaptation involve a metabolic cost, and in a natural system most plant–insect interactions involving herbivory reach a ‘stand‐off’ where both host and herbivore survive but develop suboptimally.

Contents

Summary
145

I.
Introduction
146

II.
Accumulation of defensive compounds and induced resistance
146

III.
Signalling pathways in wound‐induced resistance
147

IV.
Insect modulation of the wounding response
155

V.
Insects which evade the wounding response
156

VI.
Insect‐induced emission of volatiles and tritrophic interactions
157

VII.
Insect adaptation to plant defences
160

Conclusions
163

Acknowlegements
163

References
163
Fusarium oxysporum and its biocontrol
Authors: Fravel D. , Olivain C. , Alabouvette C.
Published: March 2003

Fusarium oxysporum is well represented among the rhizosphere microflora. While all strains exist saprophytically, some are well‐known for inducing wilt or root rots on plants whereas others are considered as nonpathogenic. Several methods based on phenotypic and genetic traits have been developed to characterize F. oxysporum strains. Results showed the great diversity affecting the soil‐borne populations of F. oxysporum. In suppressive soils, interactions between pathogenic and nonpathogenic strains result in the control of the disease. Therefore nonpathogenic strains are developed as biocontrol agents. The nonpathogenic F. oxysporum strains show several modes of action contributing to their biocontrol capacity. They are able to compete for nutrients in the soil, affecting the rate of chlamydospore germination of the pathogen. They can also compete for infection sites on the root, and can trigger plant defence reactions, inducing systemic resistance. These mechanisms are more or less important depending on the strain. The nonpathogenic F. oxysporum are easy to mass produce and formulate, but application conditions for biocontrol efficacy under field conditions have still to be determined.
Soil community feedback and the coexistence of competitors: conceptual frameworks and empirical tests
Authors: Bever James D.
Published: March 2003

A growing body of empirical work suggests that soil organisms can exert a strong role in plant community dynamics and may contribute to the coexistence of plant species. Some of this evidence comes from examining the feedback on plant growth through changes in the composition of the soil community. Host specific changes in soil community composition can generate feedback on plant growth and this feedback can be positive or negative. Previous work has demonstrated that negative soil community feedback can contribute to the coexistence of equivalent competitors. In this paper, I show that negative soil community feedback can also contribute to the coexistence of strong competitors, maintaining plant species that would not coexist in the absence of soil community dynamics. I review the evidence for soil community feedback and find accumulating evidence that soil community feedback can be common, strongly negative, and generated by a variety of complementary soil microbial mechanisms, including host‐specific changes in the composition of the rhizosphere bacteria, nematodes, pathogenic fungi, and mycorrhizal fungi. Finally, I suggest topics needing further examination.
Farmers' markets: consuming local rural produce
Authors: Trobe Helen La
Published: March 2003

Marketing food directly from producers to consumers, so circumventing the ‘middlemen’ in the food supply chain, has many potential benefits. For consumers, direct marketing initiatives are providing people with locally grown, fresh, healthy and, in many cases, organic food at affordable prices. Through buying locally grown produce, consumers are giving their support to local producers as well as helping to revitalize rural economies. Producers benefit through retaining more of the value of their produce, which can help them survive through the current crisis in UK farming. There are also environmental benefits. Creating markets where people can buy produce from local farmers and growers reduces the distance that food travels between producers and consumers, which in turn decreases global environmental pollution. One direct marketing scheme – the farmers’ market – has proved to be particularly popular with local people, producers and the local councils, organizations and institutions who are involved in setting them up. This paper focuses on one such market, the Stour Valley Farmers' Market, which commenced trading on 20th June 1999. Customers who attended the first three of these monthly markets were interviewed to investigate the reasons for their attendance at the market, and their attitudes towards a number of food issues including organic and genetically modified food, local and seasonal food and concerns they may have over the way their food is produced. The research has shown that most customers visited the markets initially out of curiosity, although some attended specifically to buy healthy fresh foods. The vast majority of interviewees expressed a preference for food which is organically grown and free from genetic modification. Organic foods are generally perceived to be healthier and more flavoursome. When buying fresh foods, interviewees stated the importance of quality and freshness in their choice of produce.
Food consumption patterns, dietary quality and health status of expectant mothers: case studies in suburban and rural communities in Ghana
Authors: Nti Christina A. , Larweh Patience M. , Gyemfua‐Yeboah Yaa
Published: March 2003

The study was conducted to determine the food consumption patterns, dietary quality and health status of expectant women and also to assess their level of awareness of nutritional requirements during pregnancy. Food beliefs, taboos and superstitions held by the women were also investigated. Using the purposive sampling technique, 30 pregnant women attending the Maternal and Child Health Clinic at Dodowa (rural) and University Hospital, Legon (suburban), were selected for the study. A structured questionnaire was used to collect information on the respondents’ nutritional knowledge, beliefs, taboos and superstitions and health status. A food frequency questionnaire and the 24‐h dietary recall method were used to obtain information on food consumption patterns and dietary quality. The study revealed that, although the majority of the respondents (83.3%) had some knowledge of nutrition, not all were applying it in their feeding practices because of financial constraints. With regard to consumption patterns, most of the women (83.3%) were having three meals a day, while the rest either ate twice a day or anytime they were hungry. Seventy‐three per cent of the respondents also increased their food intake during pregnancy. Foods eaten were based mainly on starchy roots and tubers, cereals and vegetables. Legumes, oilseeds and fruits were often lacking in the main meals of respondents. Although animal products were consumed daily, the quantities taken were very small to provide for adequate protein, especially in the rural area. Various types of food were avoided by some of the expectant mothers for reasons such as nausea, loss of appetite, taboos and superstition. With regard to dietary quality, respondents from the rural area had diets of poorer quality in terms of nutrient intake compared with those from the suburban area. In both communities, iron contents of diets were quite low. Forty‐seven per cent of the women interviewed were anaemic, with a higher prevalence of anaemia being observed in the rural community. Common ailments reported by the women were dizziness, headache, waist pains and malaria. Based on the results of the study, it was recommended that nutrition education for pregnant women should be stepped up at antenatal clinics especially in rural areas.
Cultural foodways in Sweden: repeated focus group interviews with Somalian women
Authors: Jonsson Inger M. , Hallberg Lillemor R‐M. , Gustafsson Inga‐Britt
Published: March 2003

The aim of this study was to provide increased understanding of what ‘feeding the family’ means to Somalian women in Sweden. Focus group interviews were carried out with Somalian women, analysed by means of the Grounded theory method. The results show that factors both in the family and in the outside world influenced their food choice and traditions, both in their home country of Somalia and in their new country, Sweden, after migration. The categories generated in the analysis were brought together in a model showing the women ‘struggling for their own cultural identity’ and oscillating between ‘remaining Somalian’ and ‘becoming part of Swedish society’, food being an important instrument in maintaining the cultural identity of their families. Cultural identity in this respect is a matter of the wishes of the husband, followed by those of the women and children and, at the same time, the strong pressure of the Swedish host country. This indicates how important it is for professionals to be aware of the trust people have in their own cultural food and therefore also how necessary it is to give culturally adapted food advice through public health work, in this case to Somalian families.
An assessment of ethical obligation and self‐identity in ethical consumer decision‐making: a structural equation modelling approach
Authors: Shaw Deirdre , Shiu Edward
Published: March 2003

Research has supported the addition of ethical obligation and self‐identity to models of consumer decision‐making in ‘ethical’ contexts. The particular placement of ethical obligation and self‐identity within a model of ethical consumer decision‐making remains unclear. Are these measures an antecedent to attitude or behavioural intention? This paper presents findings from a large scale survey of ethical consumers that explores, through structural equation modelling, the specific placement of these measures within a validated model of ethical consumer decision‐making, which uses the theory of planned behaviour as an initial framework. This research is examined within the ‘ethical’ context of fair trade grocery purchasing. (Fairly traded products are those purchased under equitable trading agreements, involving co‐operative rather than competitive trading principles, ensuring a fair price and fair working conditions for the producers and suppliers.)
Effect of enzymatic cross‐linking of milk proteins on functional properties of set‐style yoghurt
Authors: Lorenzen P Chr , Neve H , Mautner A , Schlimme E
Published: March 2003

This paper presents research on the effect of enzymatic cross‐linking of milk proteins on the properties of yoghurt. Whole milk was incubated with transglutaminase (TG) prior to fermentation (2 h, 40°C, E/S ratio 1/2000). Enzyme action was stopped by heating (1 min, 80°C). Skim‐milk was treated by simultaneous use of TG and thermophilic yoghurt starter culture without inactivation of the enzyme. A TG treatment of milk prior to fermentation led to prolonged fermentation, while the concomitant use of TG and culture had no influence on fermentation time. Post acidification of yoghurt during storage was lower for products from enzyme‐treated milk. This applies both for products cross‐linked prior to fermentation with enzyme inactivation, and for simultaneous use of culture and TG without inactivation of the enzyme. Scanning electron microscopic studies revealed that a TG treatment of milk led to reduced mesh sizes of the protein network, and a more regular distribution of the proteins in the yoghurt gel. As a result, yoghurt products from enzyme‐treated milk showed increased gel strength and less syneresis, especially when the enzyme was not inactivated. Sensory studies revealed that odour and consistency properties of products from TG‐treated milk were assessed as less ‘yoghurt specific’. On the other hand, products from enzyme‐treated milk were described as being more creamy, indicating that a TG treatment may simulate fat in fermented milk products.
SALUS: online co‐operation between South Australian health libraries
Authors: Peterson Mary , Harris Lindsay
Published: March 2003

The establishment of the South Australian Health Libraries Consortium led to the development of the SALUS project which enabled the online delivery of core health, clinical information and bibliographic databases with full‐text resources across South Australian government health services. This was the first venture of Australian health librarians to create an online consortium. This article discusses the policy and management issues surrounding the implementation of the SALUS project and its influence on the evolution of the health Libraries Consortium over several years. Lessons learned during this process included: the necessity to plan well ahead yet to remain flexible in implementing those plans, to avoid procedure becoming more important than improvization during planning, to be willing to take well judged risks on occasions, and to be fully aware of changing circumstances in the project’s funding and political environment. The importance of securing influential advocates outside of the participating libraries to support continued project funding is also considered.
Distance learning project—Information skills training: supporting flexible trainees in psychiatry*
Authors: Poyner Ann , Wood Aileen , Herzberg Joe
Published: March 2003

This one‐year project was designed to investigate the skills training needs of a group of 12 specialist registrars in psychiatry working as flexible trainees (part time) in the NHS. They had expressed concerns about their lack of information technology skills at training assessment seminars. A part‐time project officer, was appointed to support the group giving information skills training to each flexible trainee on an individual basis, through a series of home visits. The project officer worked under the supervision of the London Library & Information Development Unit (LLIDU) from a home base, to identify with the insular feelings expressed by the project group. Project funding purchased a laptop computer with modem and a printer for each participant to use during the project. The training covered: basic search skills and developing search strategy techniques using quality filters particularly in relation to the medline database; how to search the Cochrane Library and PubMed and exploiting Internet information resources. The group asked for help with word processing and presentation software use. These needs were met in a series of half‐day workshops. Experience and insight gained during the project will feed into future planning of information skills training for larger numbers of flexible trainees.
Molecular Cloning and Regional Distribution of a Human Proton Receptor Subunit with Biphasic Functional Properties
Authors: Babinski Kazimierz , Lê Khanh‐Tuoc , Séguéla Philippe
Published: March 2003

Abstract : Small changes of extracellular pH activate depolarizing
inward currents in most nociceptive neurons. It has been recently proposed
that acid sensitivity of sensory as well as central neurons is mediated by a
family of proton‐gated cation channels structurally related to
Caenorhabditis elegans degenerins and mammalian epithelial sodium
channels. We describe here the molecular cloning of a novel human proton
receptor, hASIC3, a 531‐amino acid‐long subunit homologous to rat DRASIC.
Expression of homomeric hASIC3 channels in Xenopus oocytes generated
biphasic inward currents elicited at pH <5, providing the first functional
evidence of a human proton‐gated ion channel. Contrary to the DRASIC current
phenotype, the fast desensitizing early component and the slow sustained late
component differed both by their cationic selectivity and by their response to
the antagonist amiloride, but not by their pH sensitivity (pH50 = 3.66 vs. 3.82). Using RT‐PCR and mRNA blot hybridization, we detected hASIC3 mRNA in sensory ganglia, brain, and many internal tissues including lung and testis, so hASIC3 gene expression was not restricted to peripheral sensory neurons. These functional and anatomical data strongly suggest that hASIC3 plays a major role in persistent proton‐induced currents occurring in physiological and pathological conditions of pH changes, likely through a tissue‐specific heteropolymerization with other members of the proton‐gated channel family.
Effects of Calyculin A and Okadaic Acid on Acetylcholine Release and Subcellular Distribution in Rat Hippocampal Formation
Authors: Issa Amalia M. , Gauthier Serge , Collier Brian
Published: March 2003

Abstract : The mechanisms regulating the compartmentation of
acetylcholine (ACh) and the relationship between transmitter release and ACh
stores are not fully understood. In the present experiments, we investigated
whether the inhibitors of serine/threonine phosphatases 1 and 2A, calyculin A
and okadaic acid, alter subcellular distribution and the release of ACh in rat
hippocampal slices. Calyculin A and okadaic acid significantly (p
< 0.05) depleted the occluded ACh of the vesicular P3 fraction,
but cytoplasmic ACh contained in the S3 fraction was not
significantly affected. The P3 fraction is known to be
heterogeneous ; calyculin A and okadaic acid reduced significantly (p
< 0.05) the amount of ACh recovered with a monodispersed fraction (D) of
synaptic vesicles, but the other nerve terminal bound pools (E‐F and G‐H) were
not so affected. K+‐evoked ACh release decreased significantly
(p < 0.01) in the presence of calyculin A and okadaic acid,
suggesting that fraction D's vesicular store of ACh contributes to transmitter
release. The loss of ACh from synaptic vesicle fractions prepared from tissue
exposed to phosphatase inhibitors appeared not to result from a reduced
ability to take up ACh. Thus, when tissue was allowed to synthesize
[3H]ACh from [3H]choline, the ratio of
[3H]ACh in the S3 to P3 fractions was not much changed by exposure of tissue to calyculin A or okadaic acid ; furthermore, the specific activity of ACh recovered from the D fraction was not reduced disproportionately to that of cytosolic ACh. The changes are considered to reflect reduced synthesis of ACh by tissue treated with the phosphatase inhibitors, rather than an effect on vesicle uptake mechanisms. Thus, exposure of tissue to calyculin A or okadaic acid appears to produce selective depletion of tissue ACh content in a subpopulation of synaptic vesicles, suggesting that phosphatases play a role in ACh compartmentation.
Blood—Brain Barrier Glucose Transporter
Authors: Simpson Ian A. , Appel Nathan M. , Hokari Mitsuhiko , Oki Jun , Holman Geoffrey D. , Maher Fran , Koehler‐Stec Ellen M. , Vannucci Susan J. , Smith Quentin R.
Published: March 2003

Abstract : The transport of glucose across the blood‐brain barrier
(BBB) is mediated by the high molecular mass (55‐kDa) isoform of the GLUT1
glucose transporter protein. In this study we have utilized the tritiated,
impermeant photolabel
2‐N‐[4‐(1‐azi‐2,2,2‐trifluoroethyl)[2‐3H]propyl]‐1,3‐bis(d‐mannose‐4‐yloxy)‐2‐propylamine
to develop a technique to specifically measure the concentration of GLUT1
glucose transporters on the luminal surface of the endothelial cells of the
BBB. We have combined this methodology with measurements of BBB glucose
transport and immunoblot analysis of isolated brain microvessels for labeled
luminal GLUT1 and total GLUT1 to reevaluate the effects of chronic
hypoglycemia and diabetic hyperglycemia on transendothelial glucose transport
in the rat. Hypoglycemia was induced with continuous‐release insulin pellets
(6 U/day) for a 12‐ to 14‐day duration ; diabetes was induced by
streptozotocin (65 mg/kg i.p.) for a 14‐ to 21‐day duration. Hypoglycemia
resulted in 25‐45% increases in regional BBB permeability‐surface area
(PA) values for d‐[14C]glucose uptake, when
measured at identical glucose concentration using the in situ brain perfusion
technique. Similarily, there was a 23 ± 4% increase in total GLUT1/mg
of microvessel protein and a 52 ± 13% increase in luminal GLUT1 in
hypoglycemic animals, suggesting that both increased GLUT1 synthesis and a
redistribution to favor luminal transporters account for the enhanced uptake.
A corresponding (twofold) increase in cortical GLUT1 mRNA was observed by in
situ hybridization. In contrast, no significant changes were observed in
regional brain glucose uptake PA, total microvessel 55‐kDa GLUT1, or luminal GLUT1 concentrations in hyperglycemic rats. There was, however, a 30‐40% increase in total cortical GLUT1 mRNA expression, with a 96% increase in the microvessels. Neither condition altered the levels of GLUT3 mRNA or protein expression. These results show that hypoglycemia, but not hyperglycemia, alters glucose transport activity at the BBB and that these changes in transport activity result from both an overall increase in total BBB GLUT1 and an increased transporter concentration at the luminal surface.
Down‐Regulation of Neuronal Nitric Oxide Synthase by Nitric Oxide After Oxygen‐Glucose Deprivation in Rat Forebrain Slices
Authors: Alba J. De , Cárdenas A. , Moro M. A. , Leza J. C. , Lorenzo P. , Boscá L. , Lizasoain I.
Published: March 2003

Abstract : The precise role that nitric oxide (NO) plays in the
mechanisms of ischemic brain damage remains to be established. The expression
of the inducible isoform (iNOS) of NO synthase (NOS) has been demonstrated not
only in blood and glial cells using in vivo models of brain
ischemia‐reperfusion but also in neurons in rat forebrain slices exposed to
oxygen‐glucose deprivation (OGD). We have used this experimental model to
study the effect of OGD on the neuronal isoform of NOS (nNOS) and iNOS. In
OGD‐exposed rat forebrain slices, a decrease in the calcium‐dependent NOS
activity was found 180 min after the OGD period, which was parallel to the
increase during this period in calcium‐independent NOS activity. Both
dexamethasone and cycloheximide, which completely inhibited the induction of
the calcium‐independent NOS activity, caused a 40‐70% recovery in
calcium‐dependent NOS activity when compared with slices collected immediately
after OGD. The NO scavenger oxyhemoglobin produced complete recovery of
calcium‐dependent NOS activity, suggesting that NO formed after OGD is
responsible for this down‐regulation. Consistently, exposure to the NO donor
(Z)‐1‐[(2‐aminoethyl)‐N‐(2‐ammonioethyl)amino]diazen‐1‐ium‐1,2‐diolate (DETA‐NONOate) for 180 min caused a decrease in the calcium‐dependent NOS activity present in control rat forebrain slices. Furthermore, OGD and DETA‐NONOate caused a decrease in level of both nNOS mRNA and protein. In summary, our results indicate that iNOS expression down‐regulates nNOS activity in rat brain slices exposed to OGD. These studies suggest important and complex interactions between NOS isoforms, the elucidation of which may provide further insights into the physiological and pathophysiological events that occur during and after cerebral ischemia.
Prostaglandin E Receptor Subtypes in Cultured Rat Microglia and Their Role in Reducing Lipopolysaccharide‐Induced Interleukin‐1 β Production
Authors: Caggiano Anthony O. , Kraig Richard P.
Published: March 2003

Abstract : Prostaglandins (PGs) are potent modulators of brain
function under normal and pathological conditions. The diverse effects of PGs
are due to the various actions of specific receptor subtypes for these
prostanoids. Recent work has shown that PGE2, while generally
considered a proinflammatory molecule, reduces microglial activation and thus
has an antiinflammatory effect on these cells. To gain further insight to the
mechanisms by which PGE2 influences the activation of microglia, we
investigated PGE receptor subtype, i.e., EP1, EP2, EP3, and EP4, expression
and function in cultured rat microglia. RT‐PCR showed the presence of the EP1
and EP2 but not EP3 and EP4 receptor subtypes. Sequencing confirmed their
identity with previously published receptor subtypes. PGE2 and the
EP1 agonist 17‐phenyl trinor PGE2 but not the EP3 agonist
sulprostone elicited reversible intracellular [Ca2+] increases in
microglia as measured by fura‐2. PGE2 and the EP2/EP4‐specific
agonists 11‐deoxy‐PGE1 and 19‐hydroxy‐PGE2 but not the
EP4‐selective agonist 1‐hydroxy‐PGE1 induced dose‐dependent
production of cyclic AMP (cAMP). Interleukin (IL)‐1β production, a marker
of activated microglia, was also measured following lipopolysaccharide
exposure in the presence or absence of the receptor subtype agonists.
PGE2 and the EP2 agonists reduced IL‐1β production. IL‐1β
production was unchanged by EP1, EP3, and EP4 agonists. The adenylyl cyclase
activator forskolin and the cAMP analogue dibutyryl cAMP also reduced
IL‐1β production. Thus, the inhibitory effects of PGE2 on
microglia are mediated by the EP2 receptor subtype, and the signaling
mechanism of this effect is likely via cAMP. These results show that the
effects of PGE2 on microglia are receptor subtype‐specific. Furthermore, they suggest that specific and selective manipulation of the effects of PGs on microglia and, as a result, brain function may be possible.
The Bacterial Endotoxin Lipopolysaccharide Causes Rapid Inappropriate Excitation in Rat Cortex
Authors: Wang Yushan S. , White Thomas D.
Published: March 2003

Abstract : There is mounting evidence that inflammation and
associated excitotoxicity may play important roles in various
neurodegenerative disorders, such as bacterial infections, Alzheimer's
disease, AIDS dementia, and multiple sclerosis. The immunogen E. coli
lipopolysaccharide (LPS, endotoxin) has been widely used to stimulate
immune/inflammatory responses both systemically and in the CNS. Here, we show
that exposure of parietal cortical slices from adult rats to LPS triggered
very rapid (<2.5 min) and sustained releases of the neurotransmitters
glutamate and noradrenaline, and of the neuromodulator adenosine. The
responses to LPS declined rapidly following removal of the LPS and exhibited
no tachyphylaxis to repeated exposures to LPS. The detoxified form of LPS had
no effect. LPS‐evoked release of [3H]noradrenaline, but not of
glutamate or adenosine, appears to be partly due to the released glutamate
acting at ionotropic receptors on the noradrenergic axons present in the
cortical slices. LPS appears to release glutamate, which then acts at non‐NMDA
receptors to remove the voltage‐sensitive Mg2+ block of NMDA receptors, thus permitting NMDA receptors to be activated and noradrenaline release to proceed. It seems possible that rapid, inappropriate excitation may occur in the immediate vicinity of gram‐negative bacterial infections in the brain. If similar inappropriate excitations are also triggered by those immunogens specifically associated with Alzheimer's disease (β‐amyloid), AIDS dementia (gp 120 and gp41), or multiple sclerosis (myelin basic protein), they might explain some of the acute, transient neurological and psychiatric symptoms associated with these disorders.
Regional Differences in Striatal Dopamine Uptake and Release Associated with Recovery from MPTP‐Induced Parkinsonism
Authors: Rothblat David S. , Schneider J. S.
Published: March 2003

Abstract : This study directly assessed striatal dopamine (DA) uptake rates and peak release in response to KCl in normal, symptomatic, and recovered 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐treated cats using in vivo electrochemistry. DA uptake rates measured after direct application of known concentrations of DA to the striatum were slowed significantly in both dorsal and ventral striatum in symptomatic cats compared with rates recorded in normal animals. DA uptake rates remained significantly slowed in recovered cats and were not significantly different from the rates recorded in symptomatic animals. In symptomatic cats, both DA uptake rates and the signal recorded in response to KCl stimulation were significantly decreased from normal in all dorsal and ventral striatal regions sampled. Reduction/oxidation (redox) ratios recorded in response to KCl stimulation suggested DA to be the predominant electroactive species. In spontaneously recovered MPTP‐treated cats, recordings in the ventral striatum subsequent to KCl stimulation again suggested DA to be the predominant electroactive species released, and peak levels were significantly higher than those recorded in symptomatic animals. In the dorsal striatum of recovered cats, redox ratios recorded subsequent to KCl stimulation suggested serotonin rather than DA to be the predominant electroactive species released. Peak levels of release in the dorsal striatum were not significantly greater than those recorded in symptomatic animals. These results suggest that in spontaneously recovered MPTP‐treated cats, there is partial recovery of ventral striatal DAergic terminals, persistent loss of dorsal striatal DAergic terminals, and a down‐regulation of DA transporter number/function throughout the striatum. These processes may contribute to volume transmission of DA in the striatum and promote functional recovery.
Dopaminergic Innervation of the Amygdala Is Highly Responsive to Stress
Authors: Inglis Fiona M. , Moghaddam Bita
Published: March 2003

Abstract : The amygdala has been implicated in the neuronal sequelae of stress, although little is known about the neurochemical mechanisms underlying amygdala transmission. In vivo microdialysis was employed to measure extracellular levels of dopamine in the basolateral nucleus of the amygdala in awake rats. Once it was established that impulse‐dependent release of dopamine could be measured reliably in the amygdala, the effect of stress, induced by mild handling, on amygdala dopamine release was compared with that in three other dopamine‐innervated regions, the medial prefrontal cortex, nucleus accumbens, and caudate nucleus. The magnitude of increase in dopamine in response to the handling stimulus was significantly greater in the amygdala than in the nucleus accumbens and prefrontal cortex. This increase was maximal during the application of stress and diminished after the cessation of stress. In contrast, the increases in extracellular dopamine levels in other regions, in particular the nucleus accumbens, were prolonged, reaching maximal values after the cessation of stress. These results suggest that dopaminergic innervation of the amygdala may be more responsive to stress than that of other dopamine‐innervated regions of the limbic system, including the prefrontal cortex, and implicate amygdalar dopamine in normal and pathophysiological processes subserving an organism's response to stress.
Blockade by NS‐7, a Neuroprotective Compound, of Both L‐Type and P/Q‐Type Ca2+ Channels Involving Depolarization‐Stimulated Nitric Oxide Synthase Activity in Primary Neuronal Culture
Authors: Oka Michiko , Itoh Yoshinori , Ukai Yojiro , Kimura Kiyoshi
Published: March 2003

Abstract : The effect of
4‐(4‐fluorophenyl)‐2‐methyl‐6‐(5‐piperidinopentyloxy)pyrimidine hydrochloride
(NS‐7), a neuroprotective compound, on Ca2+ channels involving the
activation of nitric oxide synthase (NOS) was investigated in primary neuronal
culture. The NOS activity was estimated from the cyclic GMP formation. The KCl
(25 mM)‐stimulated cyclic GMP formation was totally abolished by a
combined treatment with nifedipine and ω‐agatoxin IVA (ω‐Aga),
whereas spontaneous cyclic GMP formation was partially but significantly
reduced by nifedipine. In contrast to nifedipine, NS‐7 blocked KCl‐stimulated
cyclic GMP formation without affecting spontaneous cyclic GMP formation.
Subsequently, the effects of nifedipine and NS‐7 on L‐type Ca2+
channels were compared. Nifedipine blocked equally the cyclic GMP formation
stimulated by various concentrations of (±)‐Bay K 8644, whereas NS‐7
inhibited the maximal response without affecting the responses induced by low
concentrations of (±)‐Bay K 8644. The effects of NS‐7 on L‐type and
P/Q‐type Ca2+ channels involving KCl‐stimulated cyclic GMP
formation were subsequently examined. NS‐7 suppressed the KCl‐stimulated
cyclic GMP formation measured in the presence of ω‐Aga to almost the
same extent as that determined in the presence of nifedipine. In contrast,
NS‐7 had no influence on ionomycin‐induced enhancement of cyclic GMP
formation. Finally, NS‐7 reversed KCl‐induced elevation of the intracellular
free Ca2+ concentration. These findings suggest that NS‐7 inhibits
NOS activation in primary neuronal culture by reducing Ca2+ entry
through L‐type and P/Q‐type Ca2+ channels, in which the inhibition
is largely dependent on Ca2+ channel activity.
Synthesis, Degradation, and Subcellular Localization of Synaptotagmin IV, a Neuronal Immediate Early Gene Product
Authors: Ferguson Gregory D. , Thomas David M. , Elferink Lisa A. , Herschman Harvey R.
Published: March 2003

Abstract: Synaptotagmin IV (Syt IV) is an immediate early gene induced by depolarization in rat PC12 cells and in rat hippocampus. We prepared an antiserum to Syt IV protein. The 46‐kDa Syt IV protein is nearly undetectable by western blotting in unstimulated PC12 cells. After depolarization, Syt IV increases rapidly, peaks at 4 h, and decays to near baseline levels by 12 h. Forskolin stimulation also leads to rapid Syt IV protein accumulation. The rate of Syt IV protein synthesis, determined by labeling with radioactive amino acids and immunoprecipitation, is low in unstimulated PC12 cells, but increases over the first 3 h after forskolin stimulation and remains elevated for several hours. Syt IV protein is relatively labile; metabolically labeled Syt IV has a half‐life of ∼2 h in PC12 cells. Sucrose density gradient fractionation and vesicle immunoisolation experiments suggest that Syt IV protein is present in both synaptic‐like microvesicles and secretory granules. Vesicles immunoisolated from forskolin‐treated PC12 cells with anti‐Syt I antibody contain radioactively labeled Syt IV, demonstrating that Syt I and Syt IV colocalize in common vesicles. These results suggest that Syt IV protein, after its stimulation‐induced synthesis, is rapidly transported to secretory vesicles where it may transiently modulate the exocytotic machinery.
Regulation of Nociceptin/Orphanin FQ Gene Expression by Neuropoietic Cytokines and Neurotrophic Factors in Neurons and Astrocytes
Authors: Buzas Beata , Symes Aviva J. , Cox Brian M.
Published: March 2003

Abstract: We have identified the gene encoding nociceptin/orphanin FQ (N/OFQ), the novel opioid‐like neuropeptide, as responsive to ciliary neurotrophic factor (CNTF). N/OFQ mRNA levels were induced five‐ and ninefold by CNTF in striatal and cortical neurons. In primary astrocytes CNTF also increased N/OFQ mRNA levels. CNTF is a multifunctional cytokine that mediates the development and differentiation of both neurons and astrocytes and supports the survival of various neurons. CNTF is also an injury‐induced factor in the brain playing a crucial role in astrogliosis. The mechanism by which CNTF elicits these effects is not well understood, but it is likely to involve regulation of specific genes. CNTF regulation of N/OFQ expression was sensitive to the kinase inhibitors H‐7 and genistein but not to inhibition of protein synthesis. This pharmacological profile is consistent with CNTF activating the Janus protein tyrosine kinase (JAK)/signal transducers and activators of transcription (STAT) pathway to induce N/OFQ transcription. In nuclear extracts of CNTF‐treated striatal neurons DNA binding of STAT proteins was increased. Radioimmunoassays revealed elevated N/OFQ immunoreactivity in striatal neurons after CNTF treatment. Expression of the related proenkephalin gene was not affected by CNTF in either neuronal or glial cultures. Regulation of N/OFQ expression by CNTF might point to a possible function of N/OFQ during development and after neural injury.
Effects of Acute and Chronic Administration of the Serotonin1A Agonist Buspirone on Serotonin Synthesis in the Rat Brain
Authors: Okazawa Hidehiko , Yamane Fumitaka , Blier Pierre , Diksic Mirko
Published: March 2003

Abstract: The effects of acute and chronic administration of buspirone, a serotonin 5‐HT1A agonist, on the 5‐HT synthesis rates in various rat brain structures were investigated using α‐[14C]methyl‐L‐tryptophan (α‐[14C]MTrp) and an autoradiographic method. In the acute treatment study, buspirone (10 mg/kg) was injected subcutaneously 30 min before α‐[14C]MTrp administration (30 μCi over 2 min) into a femoral vein. In the chronic treatment study, buspirone was given in a sustained fashion (10 mg/kg/day) for 14 days using an osmotic minipump implanted subcutaneously. Rats were killed 60 and 150 min after α‐[14C]MTrp administration (two‐time point method). A single dose of buspirone induced a significant decrease of 5‐HT synthesis throughout the brain with the exception of the pineal body. However, the chronic treatment with buspirone did not induce significant differences in 5‐HT synthesis in the brain. There was no significant difference in plasma free tryptophan concentration between any of the groups. The unaltered 5‐HT synthesis rates in the chronic treatment study likely reflect a normalization of this parameter due to a desensitization of 5‐HT1A autoreceptors on the cell body of 5‐HT neurons, which has been previously shown to occur following long‐term treatment with 5‐HT1A agonists.
Exacerbation of Copper Toxicity in Primary Neuronal Cultures Depleted of Cellular Glutathione
Authors: White Anthony R. , Bush Ashley I. , Beyreuther Konrad , Masters Colin L. , Cappai Roberto
Published: March 2003

Abstract: Perturbations to glutathione (GSH) metabolism may play an important role in neurodegenerative disorders such as Alzheimer's, Parkinson's, and prion diseases. A primary function of GSH is to prevent the toxic interaction between free radicals and reactive transition metals such as copper (Cu). Due to the potential role of Cu in neurodegeneration, we examined the effect of GSH depletion on Cu toxicity in murine primary neuronal cultures. Depletion of cellular GSH with L‐buthionine‐[S,R]‐sulfoximine resulted in a dramatic potentiation of Cu toxicity in neurons without effect on iron (Fe) toxicity. Similarly, inhibition of glutathione reductase (GR) activity with 1,3‐bis(2‐chloroethyl)‐1‐nitrosurea also increased Cu toxicity in neurons. To determine if the Alzheimer's amyloid‐β (Aβ) peptide can affect neuronal resistance to transition metal toxicity, we exposed cultures to nontoxic concentrations of Aβ25‐35 in the presence or absence of Cu or Fe. Aβ25‐35 pretreatment was found to deplete neuronal GSH and increase GR activity, confirming the ability of Aβ to perturb neuronal GSH homeostasis. Aβ25‐35 pretreatment potently increased Cu toxicity but had no effect on Fe toxicity. These studies demonstrate an important role for neuronal GSH homeostasis in selective protection against Cu toxicity, a finding with widespread implications for neurodegenerative disorders.