bluuebunny/crossref_metadata_2025 huggingface dataset:
Olfactory input to the parahippocampal region of the isolated guinea pig brain reveals weak entorhinal‐to‐perirhinal interactions
Authors: Biella G. R. , Gnatkovsky V. , Takashima I. , Kajiwara R. , Iijima T. , De Curtis M.
Published: July 2003

The processing of olfactory inputs by the parahippocampal region has a central role in the organization of memory in mammals. The olfactory input is relayed to the hippocampus via interposed synapses located in the piriform and entorhinal cortices. Whether olfactory afferents directly or indirectly project to other areas of the parahippocampal region beside the entorhinal cortex (EC) is uncertain. We performed an electrophysiological and imaging study of the propagation pattern of the olfactory input carried by the fibres that form the lateral olfactory tract (LOT) into the parahippocampal region of the in vitro isolated guinea pig preparation. Laminar analysis was performed on field potential depth profiles recorded with 16‐channel silicon probes at different sites of the insular–parahippocampal cortex. The LOT input induced a large amplitude polysynaptic response in the lateral EC. Following appropriate LOT stimulation, a late response generated by the interposed activation of the hippocampus was observed in the medial EC. LOT stimulation did not induce any local response in area 36 of the perirhinal cortex (PRC), while a small amplitude potential with a delay similar to the lateral EC response was inconsistently observed in PRC area 35. No PRC potentials were observed following the responses evoked by LOT stimulation in either the lateral or the medial EC. These findings were substantiated by current source density analysis of PRC laminar profiles. To further verify the absence of EC‐to–PRC field interactions after LOT stimulation, high‐resolution optical imaging of neuronal activity was performed after perfusion of the isolated brain with the voltage‐sensitive dye RH‐795. The optical recordings confirmed that olfactory‐induced activity in the EC does not induce massive PRC activation. The present findings suggest that the olfactory input into the parahippocampal region is confined to the entorhinal cortex. The results also imply that, as demonstrated for the PRC‐to‐EC pathway, the propagation of neuronal activity from the EC to the PRC is hindered, possibly by a powerful inhibitory control generated within the EC.
Light does not degrade the constitutively expressed BMAL1 protein in the mouse suprachiasmatic nucleus
Authors: Von Gall Charlotte , Noton Elizabeth , Lee Choogon , Weaver David R.
Published: July 2003

Biological rhythms in mammals are driven by a central circadian clock located in the suprachiasmatic nucleus (SCN). At the molecular level the biological clock is based on the rhythmic expression of clock genes. Two basic helix–loop–helix (bHLH)/PAS‐containing transcription factors, CLOCK and BMAL1 (MOP3), provide the basic drive to the system by activating transcription of negative regulators through E box enhancer elements. A critical feature of circadian timing is the ability of the clockwork to be entrained to the environmental light/dark cycle. The light‐resetting mechanism of the mammalian circadian clock is poorly understood. Light‐induced phase shifts are correlated with the induction of the clock genes mPer1 and mPer2 and a subsequent increase in mPER1 protein levels. It has previously been suggested that rapid degradation of BMAL1 protein in the rat SCN is part of the resetting mechanism of the central pacemaker. Our study shows that BMAL1 and CLOCK proteins are continuously expressed at high levels in the mouse SCN, supporting the hypothesis that rhythmic negative feedback plays the major role in rhythm generation in the mammalian pacemaker. Using both immunocytochemistry and immunoblot analysis, our studies demonstrate that BMAL1 protein in the mouse SCN is not affected by a phase‐resetting light pulse. These results indicate that rapid degradation of BMAL1 protein is not a consistent feature of resetting mechanisms in rodents.
Induction of rapid, activity‐dependent neuronal–glial remodelling in the adult rat hypothalamus in vitro
Authors: Langle Sarah L. , Poulain Dominique A. , Theodosis Dionysia T.
Published: July 2003

The hypothalamic oxytocinergic system offers a remarkable model of morphological plasticity in the adult because its neurons and astrocytes undergo mutual remodelling in relation to differing physiological conditions. Among various factors involved in such plasticity, oxytocin (OT) itself appears of primary importance as its central administration resulted in morphological changes similar to those brought on by physiological stimuli. In the present study, we applied OT on acute hypothalamic slices from adult rats that included the supraoptic nucleus. Using ultrastructural morphometric analyses, we found that it induced a significant reduction of astrocytic coverage of OT neurons, leaving their surfaces directly juxtaposed, to an extent similar to that detected in vivo under conditions like lactation. These neuronal–glial changes were rapid and reversible, occurring within a few hours, and specifically mediated via OT receptors. They were potentiated by oestrogen and depended on calcium mobilization and de novo protein synthesis. Moreover, they depended on concurrent neuronal activation brought on by hyperosmotic stimulation or blockade of inhibitory GABAergic neurotransmission; they were inhibited by blockade of glutamatergic receptors. Taken together, our observations show that intrahypothalamic release of OT affects not only neuronal activation of the OT system but its morphological plasticity as well. Moreover, the activity dependence of the OT‐induced changes strongly suggests that astrocytes can sense the level of activity of adjacent neurons and/or afferent input and this can subsequently act as a signal to bring on the neuronal and glial conformational changes.
Rilmenidine produces mydriasis in cats by stimulation of CNS α2‐adrenoceptors
Authors: Koss M. C.
Published: July 2003

1
Experiments were undertaken to determine if the imidazoline/α2‐adrenoceptor agonist, rilmenidine, would produce mydriasis in cats and, if so, to delineate its site of action and determine if this effect is mediated by imidazoline receptors or α2‐adrenoceptors.

2 Rilmenidine produced dose‐related pupillary dilator responses in pentobarbital anaesthetized cats that were independent of sympathetic innervation to the iris but were dependent upon intact parasympathetic neuronal tone. The ED50 for rilmenidine‐induced pupillary dilation was approximately 200 μg kg−1, i.v., and was sustained for at least 1 h.

3 The highly selective α2‐adrenoceptor antagonist, RS‐79948, administered either before or after rilmenidine, antagonized rilmenidine‐induced mydriasis. Neuronally induced reflex inhibition of parasympathetic nerve activity was also inhibited by administration of RS‐79948.

4 These results suggest that rilmenidine acts like clonidine to produce pupillary dilation by inhibition of parasympathetic tone to the iris sphincter and that this central nervous system parasympatho‐inhibition is mediated by α2‐adrenoceptors, rather than imidazoline receptors.
Actions of domperidone on the opioid system in isolated guinea‐pig ileum: differences between dopamine antagonists on opioid system
Authors: Ozaki M. , Masuda Y. , Kishioka S. , Yamamoto H.
Published: July 2003

To validate the relationship between the dopaminergic, opioidergic and cholinergic nervous systems, we evaluated the effect of domperidone, a dopamine (D2) antagonist, on the opioid system in myenteric plexus‐longitudinal muscle preparation isolated from the guinea‐pig ileum.

One micromolar of domperidone did not affect the 0.1 Hz‐evoked (duration 0.5 ms, maximum intensity) twitch response, but concentration dependently inhibited the twitch response between concentrations of 2 and 20 μm, and the inhibition was maximum after 20–30 min at the highest concentration used (20 μm).

 Acetylcholine‐evoked contraction on basal tension was also not inhibited by 1 μm domperidone, but the contraction was concentration dependently inhibited at concentrations of 10–100 μm in a non‐competitive manner.

One micromolar of domperidone, however, increased post‐tetanic twitch inhibition, an indicator of the release of endogenous opioids. This increase was completely antagonized by 1 μm naloxone.

Twitch inhibition induced by dynorphin 1–13 (0.1–10 nm) was not affected by 1 μm domperidone, but increased the maximum twitch inhibition caused by morphine (0.1–1 μm).

These results might reflect the existence of an interaction between the dopaminergic and opioidergic system without the inhibition of the cholinergic system. Dopamine antagonists increased opioid action, an action that may depend more on the increased release of endogenous opioids than on supersensitivity of the opioid receptor.
Reflex bradycardia induced by hydralazine in sino‐aortic deafferented conscious rats
Authors: Sánchez‐Salvatori M. A. , Vidrio H.
Published: July 2003

1 It is generally recognized that the vasodilator hydralazine produces hypotension accompanied by baroreflex‐mediated tachycardia. In some experimental conditions, however, the accompanying heart rate change is bradycardia, a paradoxical response which has not been satisfactorily explained. The present study examined the possibility of hydralazine‐induced bradycardia being mediated by vagal or sympathetic afferents activated by changes in left ventricular pressure.

2 Systolic blood pressure and heart rate responses to hydralazine were recorded in conscious normotensive intact rats by a tail cuff method and compared with responses in animals subjected to previous sino‐aortic deafferentation (SAD) to remove the influence of the arterial baroreflex. Responses were also obtained after blockade of myocardial afferent vagal C‐fibres with urethane, of efferent vagal impulses to the heart with methylatropine, of positive inotropic effects of hydralazine with atenolol, and of prostanoid sensitization of myocardial nerve fibres with indomethacin.

3 Hydralazine produced hypotension and tachycardia in intact rats, and hypotension and bradycardia in SAD animals. In intact rats, this pattern was not affected by any of the pretreatments, while in SAD rats, all pretreatments reversed the bradycardia to hydralazine.

4 The present results indicate that suppression of the arterial baroreflex by SAD propitiates the appearance of a bradycardiac response to hydralazine. This reaction probably results from activation of a vagal cardiodepressant reflex originating in the heart, as suggested by its blockade by drugs acting at various sites along the reflex arch.
Increased blocking activity of combined tachykinin NK1‐ and NK2‐receptor antagonists on tachykinergic bronchomotor responses in the guinea‐pig
Authors: Corboz M. R. , Fernandez X. , Rizzo C. A. , Tozzi S. , Monahan M. E. , Hey J. A.
Published: July 2003

1 The present study compared the effect of the administration of tachykinin NK1‐ and NK2‐receptor antagonists alone and in combination on exogenous and endogenous tachykinin‐induced contractions using three different guinea‐pig airway preparations: isolated bronchus, isolated perfused lung and in vivo.

2 In the isolated bronchi, the tachykinin NK1‐receptor antagonist CP 99994 (0.01–1 μm)produced concentration‐dependent inhibition of contractions induced the tachykinin NK1‐receptor agonists substance P (SP) and [Met‐OMe11] SP ([Met‐OMe11]SP), whereas the tachykinin NK2‐receptor antagonist SR 48968 (0.1 μm) had no effect. SR 48968 (0.001–0.01 μm)concentration‐dependently inhibited contractions induced by the tachykinin NK2‐receptor agonists neurokinin A (NKA) and [β‐Ala8]‐neurokinin A (4–10) ([βAla8]‐NKA) whereas CP 99994 (0.1 μm) did not inhibit the contractions. The contractile activity of capsaicin, an agent that releases endogenous tachykinins from sensory C‐fibres, was inhibited in a concentration dependent manner by SR 48968 (0.001–0.03 μm) but not by CP 99994 (0.1 μm). Combination of CP 99994 and SR 48968 caused increased inhibitory effects on the concentration–response curves to SP, [Met‐OMe11]SP, NKA, [β‐Ala8]‐NKA and capsaicin.

3 In isolated perfused lungs, SR 48968 concentration (0.01–10 μm) dependently inhibited NKA‐, [β‐Ala8]‐NKA‐ and capsaicin‐induced bronchoconstriction whereas CP 99994 (30 μm) had no effect on SP‐, NKA‐, [β‐Ala8]‐NKA‐ and capsaicin‐induced bronchoconstriction. Combination of inactive concentrations of CP 99994 and SR 48968 produced an increased inhibitory effect on all previous stimuli‐induced bronchoconstriction.

4 In in vivo guinea‐pig studies, intravenous and oral pretreatment with SR 48968 (0.01–1 mg kg−1 i.v. and 0.1–3 mg kg−1 p.o., respectively), but not with CP 99994 (1 mg kg−1 i.v. and 0.3–30 mg kg−1 p.o., respectively), produced a dose‐dependent inhibition of the bronchoconstrictor responses induced by NKA, [β‐Ala8]‐NKA and capsaicin. CP 99994 intravenously (0.3 mg kg−1) and orally (3–10 mg kg−1) inhibited SP‐induced bronchoconstriction only. Intravenous and oral low dose combinations of CP 99994 and SR 48968 produced an increased inhibition of SP‐, NKA‐, [β‐Ala8]‐NKA‐ and capsaicin‐induced bronchoconstriction, respectively.

5 The present data indicate that combined tachykinin NK1‐ and NK2‐receptor antagonist treatment compared with single antagonist treatment, using CP 99994 and SR 48968, produced an augmented blockade of tachykinin NK1‐, NK2‐ and capsaicin‐mediated contractions in guinea pig airways. These findings support the hypothesis that a dual NK1‐ and NK2‐receptor antagonist may provide an advantage over single activity tachykinin NK1‐ or NK2‐receptor antagonists in pulmonary obstructive diseases.
The effect of chronic ethanol administration on nitric oxide‐mediated responses in rat isolated trachea preparation
Authors: Usta C. , Şadan G.
Published: July 2003

1 In the present study, we investigated the effect of chronic ethanol administration on nitric oxide (NO)‐mediated responses in rat isolated trachea preparation.

2 Ethanol was given to rats in a modified liquid diet for 21 days. Isolated tracheal rings were then used to obtain responses to electrical field stimulation (EFS) after precontraction with 100 μm histamine. The parameters of field stimulation were as follows: supramaximal voltage of 50 V, 0.5 ms duration, 10‐s train; 0.5, 1, 3, 5, 10, 20, 30 and 50 Hz at 2‐min intervals. The effects of l‐and d‐arginine (10−6 m) on the responses to field stimulation (10–20 Hz) were studied. In other experiments, we tested the effects of Nω‐nitro‐l‐arginine methyl ester (l‐NAME, 10−6–10−5 m) and SIN‐1 (10−6–10−5 m) on the responses to field stimulation.

3 Electrical field stimulation induced relaxation responses in the tracheal rings precontracted with histamine from control‐ and ethanol‐treated rats. The relaxation responses induced by EFS were significantly reduced in the tracheal rings precontracted with histamine from ethanol dependent group. The responses induced by EFS in both groups were completely abolished by tetrodotoxin (1 μm), but unaffected by hexamethonium (1 μm). Incubation with d‐arginine did not cause statistically significant increases in relaxation responses to EFS in both groups. l‐Arginine (10−6 m) caused statistically significant increases in relaxation responses to EFS in control rats, but not in ethanol dependent rats. Incubation with l‐NAME (10−6–10−5 m) caused statistically significant inhibition of the relaxation responses to EFS in both groups. SIN‐1 (10−6–10−5 m) induced significantly increase in relaxation responses to EFS in both groups.

4 Our results suggest that the possible mechanism responsible for inhibition of tracheal inhibitory responses to EFS in ethanol‐dependent rats may be a reduction in production of NO and in the uptake of l‐arginine.
Inhibition of calcium/calmodulin‐dependent protein kinase II normalizes diabetes‐induced abnormal vascular reactivity in the rat perfused mesenteric vascular bed
Authors: Yousif M. H. M. , Benter I. F. , Akhtar S.
Published: July 2003

1 Calcium/calmodulin‐dependent protein kinase II (CaMKII) has an important function in mediating insulin release but its role in the development of diabetes‐induced cardiovascular complications is not known.

2 We investigated the ability of a chronic administration of KN‐93 (5 mg kg−1alt diem for 4 weeks), an inhibitor of CaMKII, to modulate the altered vasoreactivity of the perfused mesenteric bed to common vasoconstrictors and vasodilators in streptozotocin (STZ)‐induced diabetes.

3 The vasoconstrictor responses induced by noradrenaline (NE), endothelin‐1 (ET‐1), and angiotensin II (Ang II), were significantly increased whereas, vasodilator responses to carbachol and histamine were significantly reduced in the perfused mesenteric bed of the STZ‐diabetic rats as compared with non‐diabetic controls.

4 Inhibition of CaMKII by KN‐93 treatment did not affect blood glucose levels but produced a significant normalization of the altered agonist‐induced vasoconstrictor and vasodilator responses. KN‐93 did not affect agonist‐induced responses in control animals. In addition, KN‐93 significantly reduced weight loss in diabetic rats.

5 The present data suggest that CaMKII is an essential mediator in the development of diabetic vascular dysfunction and may also play an important role in signalling pathways leading to weight loss during diabetes.
The Added Clinical Value of Second Generation Ultrasound Contrast Agents
Authors: Kuecherer Helmut
Published: July 2003

Intravenous contrast echocardiography using second generation contrast agents improves wall motion analysis in difficult‐to‐image patients and increases diagnostic accuracy of stress echocardiography. New and still experimental technology enables simultaneous evaluation of myocardial perfusion and wall motion. First clinical and experimental studies demonstrate an acceptable diagnostic accuracy of intravenous contrast echocardiography in detecting stress‐induced ischemia and in differentiating viable from irreversibly damaged myocardium. Current data encourage the use of intravenous contrast echocardiography to evaluate microvascular integrity in patients with unstable angina and in acute infarction following coronary revascularization. However, attenuation artefacts may limit interpretation of findings in basal myocardial segments. Standardization of the method on the basis of more solid data derived from large clinical trials is needed before introducing intravenous myocardial contrast echocardiography into clinical practice. (ECHOCARDIOGRAPHY, Volume 20, Supplement 1, 2003)
Clinical Benefits of Using Second Generation Ultrasound Contrast Agents in Stress Echocardiography
Authors: Jung Philip H. , Angermann C. E.
Published: July 2003

Stress echocardiography is an established and widely used method for the noninvasive detection of myocardial ischemia. However, despite introduction of new echocardiographic technologies, such as harmonic imaging, unsatisfactory image quality limits an accurate interpretation of left ventricular (LV) wall thickening and motion in up to 30% of patients during stress echocardiography. Development and availability of second generation transpulmonary contrast agents, which opacify the LV chamber and have the capability of enhancing endocardial border definition, facilitate high‐quality imaging of LV structures even in technically difficult patients. Application of a contrast agent is not associated with significant extra cost of time and manipulations because bolus injections are sufficient in most instances and harmonic imaging capabilities as well as contrast specific presets are implemented and, thus, readily available in most modern ultrasound systems. Numerous studies have demonstrated that contrast echocardiography substantially improves LV cavity visualization at rest as well as at peak stress and, therefore, increases reader confidence and decreases interobserver variability. Moreover, enhanced learning curves for interpreting stress echocardiograms have been reported in novice readers. It has been reported that compared with native stress echocardiography the use of contrast results in identification of more true positive as well as true negative results and helps to avoid unnecessary invasive procedures in a considerable number of patients. Contrast stress echocardiography may, therefore, also prove cost effective in the future. Further refinements of contrast agent properties and new developments in imaging technology will likely continue to extend the spectrum of diagnostic cardiac imaging techniques and further enhance noninvasive assessment of the complex pathophysiology of coronary artery disease. (ECHOCARDIOGRAPHY, Volume 20, Supplement 1, 2003)
Assessment of Myocardial Viability in Patients With Postischemic Left Ventricular Dysfunction: Role of Myocardial Contrast Echocardiography
Authors: Agati Luciano , Majo Francesca De , Madonna Maria Pina , Celani Flavia , Funaro Stefania , Tonti Gianni
Published: July 2003

The distinction between viable and nonviable dysfunctional left ventricular (LV) segments after acute myocardial infarction is very important, because revascularization increases survival only in patients with viable myocardial tissue. Recent studies have highlighted a mismatch between two highly specific investigations for viability assessment: dobutamine echocardiography, which measures inotropic reserve, and myocardial contrast echocardiography (MCE), which measures microvascular perfusion. Viability and functional reserve are not synonymous. Maintenance of microvascular perfusion, independently of functional reserve, attenuates left ventricular remodelling, reduces the risk of major cardiac events, and increases survival. MCE provides similar perfusion information as myocardial blush, but image quality is much higher. Quantitative analysis of digital data provides more accurate diagnostic MCE information than qualitative analysis of video signal intensity. In a recent study relating MCE findings to histologic data, MCE‐derived quantitative data were closely correlated with microvascular density and capillary area, and inversely correlated with collagen content. One of the contrast agents routinely used for MCE is SonoVue, a second generation microbubble contrast agent, which is characterized by high response to ultrasound energy, ease of destruction at high energy, and strong harmonic signal at low energy. Recommendations for the assessment of postischemic LV dysfunction: routine use of MCE, followed by dobutamine echocardiography if perfusion is documented. If MCE is negative, revascularization is not indicated; if both tests are positive, revascularization is strongly recommended; if they are discordant, useful information can be obtained by assessing the extent of 201T1 viability. (ECHOCARDIOGRAPHY, Volume 20, Supplement 1, 2003)
Optimal Use of Contrast Echo in Daily Practice: What Do We Need?
Authors: Becher Harald , Timperley Jonathan
Published: July 2003

With new low power imaging and contrast application technology, it seems possible for the first time to develop a protocol that combines acquisition of both function and perfusion data using a simple modality. For this purpose a European network has been created and this group is currently developing a protocol using a commercially available ultrasound scanner and the infusion of the echocontrast agent SonoVue® (Bracco). This study describes the technical developments that will promote a comprehensive clinical contrast protocol for assessment of patients with ischemic heart disease. (ECHOCARDIOGRAPHY, Volume 20, Supplement 1, 2003)
Rpb4p, a Subunit of RNA Polymerase II, Mediates mRNA Export during Stress
Authors: Farago Marganit , Nahari Tal , Hammel Christopher , Cole Charles N. , Choder Mordechai
Published: July 2003

Changes in gene expression represent a major mechanism by which cells respond to stress. We and other investigators have previously shown that the yeast RNA polymerase II subunit Rpb4p is required for transcription under various stress conditions, but not under optimal growth conditions. Here we show that, in addition to its role in transcription, Rpb4p is also required for mRNA export, but only when cells are exposed to stress conditions. The roles of Rpb4p in transcription and in mRNA export can be uncoupled genetically by specific mutations in Rpb4p. Both functions of Rpb4p are required to maintain cell viability during stress. We propose that Rpb4p participates in the cellular responses to stress at the interface of the transcription and the export machineries.
Inducible Systemic RNA Silencing inCaenorhabditis elegans
Authors: Timmons Lisa , Tabara Hiroaki , Mello Craig C. , Fire Andrew Z.
Published: July 2003

Introduction of double-stranded RNA (dsRNA) can elicit a gene-specific RNA interference response in a variety of organisms and cell types. In many cases, this response has a systemic character in that silencing of gene expression is observed in cells distal from the site of dsRNA delivery. The molecular mechanisms underlying the mobile nature of RNA silencing are unknown. For example, although cellular entry of dsRNA is possible, cellular exit of dsRNA from normal animal cells has not been directly observed. We provide evidence that transgenic strains of Caenorhabditis elegans transcribing dsRNA from a tissue-specific promoter do not exhibit comprehensive systemic RNA interference phenotypes. In these same animals, modifications of environmental conditions can result in more robust systemic RNA silencing. Additionally, we find that genetic mutations can influence the systemic character of RNA silencing in C. elegans and can separate mechanisms underlying systemic RNA silencing into tissue-specific components. These data suggest that trafficking of RNA silencing signals in C. elegans is regulated by specific physiological and genetic factors.
Temperature Shift Coordinately Changes the Activity and the Methylation State of Transposon Tam3 in Antirrhinum majus
Authors: Hashida Shin-nosuke , Kitamura Ken , Mikami Tetsuo , Kishima Yuji
Published: July 2003

The transposition frequency of Tam3 in Antirrhinum majus, unlike that of most other cut-and-paste-type transposons, is tightly controlled by temperature: Tam3 transposes rarely at 25°C, but much more frequently at 15°C. Here, we studied the mechanism of the low-temperature-dependent transposition (LTDT) of Tam3. Our results strongly suggest that LTDT is not likely to be due to either transcriptional regulation or posttranscriptional regulation of the Tam3 TPase gene. We found that temperature shift induced a remarkable change of the methylation state unique to Tam3 sequences in the genome: Higher temperature resulted in hypermethylation, whereas lower temperature resulted in reduced methylation. The methylation state was reversible within a single generation in response to a temperature shift. Although our data demonstrate a close link between LTDT and the methylation of Tam3, they also suggest that secondary factor(s) other than DNA methylation is involved in repression of Tam3 transposition.
Cytokinin Oxidase Gene Expression in Maize Is Localized to the Vasculature, and Is Induced by Cytokinins, Abscisic Acid, and Abiotic Stress
Authors: Brugière Norbert , Jiao Shuping , Hantke Sabine , Zinselmeier Chris , Roessler Jeffrey A. , Niu Xiaomu , Jones Robert J. , Habben Jeffrey E.
Published: July 2003

Cytokinins are hormones that play an essential role in plant growth and development. The irreversible degradation of cytokinins, catalyzed by cytokinin oxidase, is an important mechanism by which plants modulate their cytokinin levels. Cytokinin oxidase has been well characterized biochemically, but its regulation at the molecular level is not well understood. We isolated a cytokinin oxidase open reading frame from maize (Zea mays), called Ckx1, and we used it as a probe in northern and in situ hybridization experiments. We found that the gene is expressed in a developmental manner in the kernel, which correlates with cytokinin levels and cytokinin oxidase activity. In situ hybridization with Ckx1 and transgenic expression of a transcriptional fusion of the Ckx1 promoter to the Escherichia coli β-glucuronidase reporter gene revealed that the gene is expressed in the vascular bundles of kernels, seedling roots, and coleoptiles. We show that Ckx1 gene expression is inducible in various organs by synthetic and natural cytokinins. Ckx1 is also induced by abscisic acid, which may control cytokinin oxidase expression in the kernel under abiotic stress. We hypothesize that under non-stress conditions, cytokinin oxidase in maize plays a role in controlling growth and development via regulation of cytokinin levels transiting in the xylem. In addition, we suggest that under environmental stress conditions, cytokinin oxidase gene induction by abscisic acid results in aberrant degradation of cytokinins therefore impairing normal development.
Computational Approaches to Identify Promoters and cis-Regulatory Elements in Plant Genomes
Authors: Rombauts Stephane , Florquin Kobe , Lescot Magali , Marchal Kathleen , Rouzé Pierre , Van de Peer Yves
Published: July 2003

The identification of promoters and their regulatory elements is one of the major challenges in bioinformatics and integrates comparative, structural, and functional genomics. Many different approaches have been developed to detect conserved motifs in a set of genes that are either coregulated or orthologous. However, although recent approaches seem promising, in general, unambiguous identification of regulatory elements is not straightforward. The delineation of promoters is even harder, due to its complex nature, and in silico promoter prediction is still in its infancy. Here, we review the different approaches that have been developed for identifying promoters and their regulatory elements. We discuss the detection of cis-acting regulatory elements using word-counting or probabilistic methods (so-called “search by signal” methods) and the delineation of promoters by considering both sequence content and structural features (“search by content” methods). As an example of search by content, we explored in greater detail the association of promoters with CpG islands. However, due to differences in sequence content, the parameters used to detect CpG islands in humans and other vertebrates cannot be used for plants. Therefore, a preliminary attempt was made to define parameters that could possibly define CpG and CpNpG islands in Arabidopsis, by exploring the compositional landscape around the transcriptional start site. To this end, a data set of more than 5,000 gene sequences was built, including the promoter region, the 5′-untranslated region, and the first introns and coding exons. Preliminary analysis shows that promoter location based on the detection of potential CpG/CpNpG islands in the Arabidopsis genome is not straightforward. Nevertheless, because the landscape of CpG/CpNpG islands differs considerably between promoters and introns on the one side and exons (whether coding or not) on the other, more sophisticated approaches can probably be developed for the successful detection of “putative” CpG and CpNpG islands in plants.
Calmodulin Is Involved in Heat Shock Signal Transduction in Wheat
Authors: Liu Hong-Tao , Li Bing , Shang Zhong-Lin , Li Xiao-Zhi , Mu Rui-Ling , Sun Da-Ye , Zhou Ren-Gang
Published: July 2003

The involvement of calcium and calcium-activated calmodulin (Ca2+-CaM) in heat shock (HS) signal transduction in wheat (Triticum aestivum) was investigated. Using Fluo-3/acetoxymethyl esters and laser scanning confocal microscopy, it was found that the increase of intracellular free calcium ion concentration started within 1 min after a 37°C HS. The levels of CaM mRNA and protein increased during HS at 37°C in the presence of Ca2+. The expression of hsp26 and hsp70 genes was up-regulated by the addition of CaCl2 and down-regulated by the calcium ion chelator EGTA, the calcium ion channel blockers LaCl3 and verapamil, or the CaM antagonists N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide and chlorpromazine. Treatment with Ca2+ also increased, and with EGTA, verapamil, chlorpromazine, or trifluoperazine decreased, synthesis of HS proteins. The temporal expression of the CaM1-2 gene and the hsp26 and hsp70 genes demonstrated that up-regulation of the CaM1-2 gene occurred at 10 min after HS at 37°C, whereas that of hsp26 and hsp70 appeared at 20 min after HS. A 5-min HS induced expression of hsp26 after a period of recovery at 22°C after HS at 37°C. Taken together, these results indicate that Ca2+-CaM is directly involved in the HS signal transduction pathway. A working hypothesis about the relationship between upstream and downstream of HS signal transduction is presented.
Desiccation Tolerance of Five Tropical Seedlings in Panama. Relationship to a Field Assessment of Drought Performance
Authors: Tyree Melvin T. , Engelbrecht Bettina M.J. , Vargas Gustavo , Kursar Thomas A.
Published: July 2003

Studies of the desiccation tolerance of the seedlings of five tropical trees were made on potted plants growing in a greenhouse. Pots were watered to field capacity and then dehydrated for 3 to 9 weeks to reach various visual wilting stages, from slightly wilted to dead. Saturated root hydraulic conductance was measured with a high-pressure flowmeter, and whole-stem hydraulic conductance was measured by a vacuum chamber method. Leaf punches (5.6-mm diameter) were harvested for measurement of leaf water potential by a thermocouple psychrometer method and for measurement of fresh and dry weight. In a parallel study, the same five species were studied in a field experiment in the understory of a tropical forest, where these species frequently germinate. Control seedlings were maintained in irrigated plots during a dry season, and experimental plants were grown in similar plots with rain exclusion shelters. Every 2 to 4 weeks, the seedlings were scored for wilt state and survivorship. After a 22-week drought, the dry plots were irrigated for several weeks to verify visual symptoms of death. The field trials were used to rank drought performance of species, and the greenhouse desiccation studies were used to determine the conditions of moribund plants. Our conclusion is that the desiccation tolerance of moribund plants correlated with field assessment of drought-performance for the five species (r  2 > 0.94).
PINOID-Mediated Signaling Involves Calcium-Binding Proteins
Authors: Benjamins René , Ampudia Carlos S. Galván , Hooykaas Paul J.J. , Offringa Remko
Published: July 2003

The plant hormone auxin is a central regulator of plant development. In Arabidopsis, the PINOID (PID) protein serine/threonine kinase is a key component in the signaling of this phytohormone. To further investigate the biological function of PID, we performed a screen for PID-interacting proteins using the yeast two-hybrid system. Here, we show that PID interacts with two calcium-binding proteins: TOUCH3 (TCH3), a calmodulin-related protein, and PID-BINDING PROTEIN 1 (PBP1), a previously uncharacterized protein containing putative EF-hand calcium-binding motifs. The interaction between PID and the calcium-binding proteins is significant because it is calcium dependent and requires an intact PID protein. Furthermore, the expression of all three genes (PID, TCH3, and PBP1) is up-regulated by auxin. TCH3 and PBP1 are not targets for phosphorylation by PID, suggesting that these proteins act upstream of PID. PBP1 was found to stimulate the autophosphorylation activity of PID, and calcium influx and calmodulin inhibitors where found to enhance the activity of PID in vivo. Our results indicate that TCH3 and PBP1 interact with the PID protein kinase and regulate the activity of this protein in response to changes in calcium levels. This work provides the first molecular evidence for the involvement of calcium in auxin-regulated plant development.
C4 Photosynthesis at Low Temperature. A Study Using Transgenic Plants with Reduced Amounts of Rubisco
Authors: Kubien David S. , von Caemmerer Susanne , Furbank Robert T. , Sage Rowan F.
Published: July 2003

C4 plants are rare in the cool climates characteristic of high latitudes and elevations, but the reasons for this are unclear. We tested the hypothesis that CO2 fixation by Rubisco is the rate-limiting step during C4 photosynthesis at cool temperatures. We measured photosynthesis and chlorophyll fluorescence from 6°C to 40°C, and in vitro Rubisco and phosphoenolpyruvate carboxylase activity from 0°C to 42°C, in Flaveria bidentis modified by an antisense construct (targeted to the nuclear-encoded small subunit of Rubisco, anti-RbcS) to have 49% and 32% of the wild-type Rubisco content. Photosynthesis was reduced at all temperatures in the anti-Rbcs plants, but the thermal optimum for photosynthesis (35°C) did not differ. The in vitro turnover rate (kcat) of fully carbamylated Rubisco was 3.8 mol mol–  1 s–  1 at 24°C, regardless of genotype. The in vitro kcat (Rubisco Vcmax per catalytic site) and in vivo kcat (gross photosynthesis per Rubisco catalytic site) were the same below 20°C, but at warmer temperatures, the in vitro capacity of the enzyme exceeded the realized rate of photosynthesis. The quantum requirement of CO2 assimilation increased below 25°C in all genotypes, suggesting greater leakage of CO2 from the bundle sheath. The Rubisco flux control coefficient was 0.68 at the thermal optimum and increased to 0.99 at 6°C. Our results thus demonstrate that Rubisco capacity is a principle control over the rate of C4 photosynthesis at low temperatures. On the basis of these results, we propose that the lack of C4 success in cool climates reflects a constraint imposed by having less Rubisco than their C3 competitors.
A New Algorithm for Computational Image Analysis of Deformable Motion at High Spatial and Temporal Resolution Applied to Root Growth. Roughly Uniform Elongation in the Meristem and Also, after an Abrupt Acceleration, in the Elongation Zone
Authors: van der Weele Corine M. , Jiang Hai S. , Palaniappan Krishnan K. , Ivanov Viktor B. , Palaniappan Kannappan , Baskin Tobias I.
Published: July 2003

A requirement for understanding morphogenesis is being able to quantify expansion at the cellular scale. Here, we present new software (RootflowRT) for measuring the expansion profile of a growing root at high spatial and temporal resolution. The software implements an image processing algorithm using a novel combination of optical flow methods for deformable motion. The algorithm operates on a stack of nine images with a given time interval between each (usually 10 s) and quantifies velocity confidently at most pixels of the image. The root does not need to be marked. The software calculates components of motion parallel and perpendicular to the local tangent of the root's midline. A variation of the software has been developed that reports the overall root growth rate versus time. Using this software, we find that the growth zone of the root can be divided into two distinct regions, an apical region where the rate of motion, i.e. velocity, rises gradually with position and a subapical region where velocity rises steeply with position. In both zones, velocity increases almost linearly with position, and the transition between zones is abrupt. We observed this pattern for roots of Arabidopsis, tomato (Lycopersicon lycopersicum), lettuce (Lactuca sativa), alyssum (Aurinia saxatilis), and timothy (Phleum pratense). These velocity profiles imply that relative elongation rate is regulated in a step-wise fashion, being low but roughly uniform within the meristem and then becoming high, but again roughly uniform, within the zone of elongation. The executable code for RootflowRT is available from the corresponding author on request.
Glycosylation Motifs That Direct Arabinogalactan Addition to Arabinogalactan-Proteins
Authors: Tan Li , Leykam Joseph F. , Kieliszewski Marcia J.
Published: July 2003

Hydroxyproline (Hyp)-rich glycoproteins (HRGPs) participate in all aspects of plant growth and development. HRGPs are generally highly O-glycosylated through the Hyp residues, which means carbohydrates help define the interactive molecular surface and, hence, HRGP function. The Hyp contiguity hypothesis predicts that contiguous Hyp residues are sites of HRGP arabinosylation, whereas clustered noncontiguous Hyp residues are sites of galactosylation, giving rise to the arabinogalactan heteropolysaccharides that characterize the arabinogalactan-proteins. Early tests of the hypothesis using synthetic genes encoding only clustered noncontiguous Hyp in the sequence (serine [Ser]-Hyp-Ser-Hyp)n or contiguous Hyp in the series (Ser-Hyp-Hyp)n and (Ser-Hyp-Hyp-Hyp-Hyp)n confirmed that arabinogalactan polysaccharide was added only to noncontiguous Hyp, whereas arabinosylation occurred on contiguous Hyp. Here, we extended our tests of the codes that direct arabinogalactan polysaccharide addition to Hyp by building genes encoding the repetitive sequences (alanine [Ala]-proline [Pro]-Ala-Pro)n, (threonine [Thr]-Pro-Thr-Pro)n, and (valine [Val]-Pro-Val-Pro)n, and expressing them in tobacco (Nicotiana tabacum) Bright-Yellow 2 cells as fusion proteins with green fluorescent protein. All of the Pro residues in the (Ala-Pro-Ala-Pro)n fusion protein were hydroxylated and consistent with the hypothesis that every Hyp residue was glycosylated with arabinogalactan polysaccharide. In contrast, 20% to 30% of Pro residues remained non-hydroxylated in the (Thr-Pro-Thr-Pro)n, and (Val-Pro-Val-Pro)n fusion proteins. Furthermore, although 50% to 60% of the Hyp residues were glycosylated with arabinogalactan polysaccharide, some remained non-glycosylated or were arabinosylated. These results suggest that the amino acid side chains of flanking residues influence the extent of Pro hydroxylation and Hyp glycosylation and may explain why isolated noncontiguous Hyp in extensins do not acquire an arabinogalactan polysaccharide but are arabinosylated or remain non-glycosylated.
Expression of Anthocyanins and Proanthocyanidins after Transformation of Alfalfa with Maize Lc
Authors: Ray Heather , Yu Min , Auser Patricia , Blahut-Beatty Laureen , McKersie Brian , Bowley Steve , Westcott Neil , Coulman Bruce , Lloyd Alan , Gruber Margaret Y.
Published: July 2003

Three anthocyanin regulatory genes of maize (Zea mays; Lc, B-Peru, and C1) were introduced into alfalfa (Medicago sativa) in a strategy designed to stimulate the flavonoid pathway and alter the composition of flavonoids produced in forage. Lc constructs included a full-length gene and a gene with a shortened 5′-untranslated region. Lc RNA was strongly expressed in Lc transgenic alfalfa foliage, but accumulation of red-purple anthocyanin was observed only under conditions of high light intensity or low temperature. These stress conditions induced chalcone synthase and flavanone 3-hydroxylase expression in Lc transgenic alfalfa foliage compared with non-transformed plants. Genotypes containing the Lc transgene construct with a full-length 5′-untranslated region responded more quickly to stress conditions and with a more extreme phenotype. High-performance liquid chromatography analysis of field-grown tissue indicated that flavone content was reduced in forage of the Lc transgenic plants. Leucocyanidin reductase, the enzyme that controls entry of metabolites into the proanthocyanidin pathway, was activated both in foliage and in developing seeds of the Lc transgenic alfalfa genotypes. Proanthocyanidin polymer was accumulated in the forage, but (+)-catechin monomers were not detected. B-Peru transgenic and C1 transgenic populations displayed no visible phenotypic changes, although these transgenes were expressed at detectable levels. These results support the emerging picture of Lc transgene-specific patterns of expression in different recipient species. These results demonstrate that proanthocyanidin biosynthesis can be stimulated in alfalfa forage using an myc-like transgene, and they pave the way for the development of high quality, bloat-safe cultivars with ruminal protein bypass.
Emerging production systems in the transnationalisation of German car manufacturers: adaptation, application or innovation?
Authors: Pries Ludger
Published: July 2003

Based on case studies, this article seeks to determine whether or not German car manufacturers are exporting a ‘German production model’ to overseas plants and to what extent the overseas operations have a reciprocal effect on production methods in the originating country. It is argued that the most adequate analytical concept is one that emphasises interest‐driven organisational learning processes.
Integrating the supply chain: industrial relations implications in US grocery distribution
Authors: Lund John , Wright Christopher
Published: July 2003

Recent technological innovations have emphasised increasing integration between enterprises across the supply chain. This article reviews these developments within US grocery distribution and highlights the potential for supply chain integration to increase union bargaining power, as well as employer strategies aimed at reducing the threat of industrial vulnerability resulting from cross‐enterprise interdependence.
A failed panopticon: surveillance of nursing practice via new technology
Authors: Timmons Stephen
Published: July 2003

New technology being introduced into nursing makes possible the surveillance of practice via an ‘electronic panopticon’. This paper reports on a qualitative study of these systems in the UK NHS, where the attempt to monitor nursing practice failed, not least due to the complexities of nursing culture and practice at ward level.
Price Differentials between Dual‐class Stocks: Voting Premium or Liquidity Discount?
Authors: Neumann Robert
Published: July 2003

A series of papers suggest that private benefits can explain the price differentials between stock classes carrying different voting rights. However, in Denmark the premium is negative for several firms over long periods. This indicates that in the absence of takeover contests, where the voting right becomes crucial in a transfer of corporate control, the price differential in stock classes with identical dividend rights is more likely to reflect investors’ liquidity risks. Whereas the existing literature tends to focus primarily on corporate control‐related explanations, this paper documents the impact of liquidity on price spreads between dual‐class shares.
Firm Defaults and the Correlation Effect
Authors: Gersbach Hans , Lipponer Alexander
Published: July 2003

We examine how the correlations of bank loan defaults depend on the correlations of asset returns and how correlations and diversification are affected by macroeconomic risks. We highlight the main properties of the relationship between asset returns and default correlations, illustrating how adverse macroeconomic shocks raise not only the likelihood of defaults, but also the correlation of defaults. The latter effect, called correlation effect, may account for more than 50% of the increase in the credit risk.
Accounting for Accountability and Management in NPOs. A Comparative Study of Four Countries: Canada, the United Kingdom, the USA and Spain
Authors: Torres Lourdes , Pina Vicente
Published: July 2003

Financial and non–financial information are developing issues in the NPO field. Countries such as Canada, the UK, the USA and Spain have recently updated their accounting systems for NPOs through the implementation of full accrual basis to enhance their accountability and the usefulness of accounting information for decision–making purposes. The information provided by accrual accounting will be incomplete until performance indicators are developed. The performance indicators are essential for making budgets, for planning and forecasting, for evaluating the financial needs, for carrying out benchmarking with other NPOs or governmental entities, and for explaining the welfare activities realised to donors.