bluuebunny/crossref_metadata_2025 huggingface dataset:
The interaction of surface plasmon polaritons with a silver film edge
Authors: Dawson P. , Puygranier B. A. F. , Cao W. , De Fornel F.
Published: March 2003

A prism coupling arrangement is used to excite surface plasmons at the surface of a thin silver film and a photon scanning tunnelling microscope is used to detect the evane‐scent field above the silver surface. Excitation of the silver/air mode of interest is performed at λ1 = 632.8 nm using a tightly focused beam, while the control of the tip is effected by exciting a counter‐propagating surface plasmon field at a different wavelength, λ2 = 543.5 nm, using an unfocused beam covering a macroscopic area. Propagation of the red surface plasmon is evidenced by an exponential tail extending away from the launch site, but this feature is abruptly truncated if the surface plasmon encounters the edge of the silver film — there is no specularly reflected ‘beam’. Importantly, the radiative decay of the surface mode at the film edge is observable only at larger tip–sample separations, emphasizing the importance of accessing the mesoscopic regime.
Effect of anticoagulation and lavage prior to casting of postmortem material with Mercox® and Batson® 17
Authors: Martín‐Orti  , Stefanov  , Gaspar  , Martín  , Martín‐Alguacil
Published: March 2003

In this study we compare the quality of vascular casts, obtained from organs of several animal species from different sources and sacrified under different conditions. Organs from healthy animals were injected with two different polymers such as Mercox and Batson No. 17. When the specimens were observed under a scanning electron microscope structural elements such as endothelial nuclear impressions on vessels and capillaries, endothelial cell borders, venous valves, imprints of smooth muscle cells and intra‐arterial cushions were identified. Organs excised post mortem from large animals can be used for microvascular corrosion casting studies with optimal results.
Imaging of various surface properties of fluorescently labelled phospholipid Langmuir–Blodgett films with a combined scanning probe microscope
Authors: Horiuchi Y. , Yagi K. , Hosokawa T. , Yamamoto N. , Muramatsu H. , Fujihira M.
Published: March 2003

We present results of phase separation of a single‐component system of 1,2‐dihexadecanoyl‐sn‐glycero‐3‐phospho‐[N‐(4‐nitrobenz)‐2‐oxa‐1,3‐diazolyl]ethanolamine in which a liquid‐condensed (LC) phase co‐exists with a liquid‐expanded (LE) phase. Domain formation in the co‐existence region was studied using a newly developed combined scanning near‐field optical microscope–atomic force microscope (SNOM–AFM). We demonstrate for the first time that the topographic, friction, fluorescence and surface potential distributions for a phase‐separated single‐component Langmuir–Blodgett film between the LE and LC phases can be simultaneously observed using the SNOM–AFM with a thin‐step etched optical fibre probe.
Stereoscopic display of atomic force microscope images using anaglyph techniques
Authors: Smith  , Connell  , Swift
Published: March 2003

This paper describes the use of a standard stereo‐pair image display method for presenting the three‐dimensional relief information found in atomic force microscope (AFM) images. The method makes use of commercially available image processing software packages. The techniques are illustrated on AFM images of the cuticle structure of a human hair fibre.
Analysis of SiC fibres and composites using Raman microscopy
Authors: Young  , Broadbridge  , So
Published: March 2003

The application of Raman spectroscopy in the analysis of the microstructure of SCS‐6 silicon carbide fibres using a Renishaw Raman microscope is described. It is demonstrated that the technique allows a detailed study to be made of the point‐to‐point variation in microstructure across a fibre section. It has been possible to monitor the variation of the concentration of SiC and carbon in the fibre microstructure and to detect differences in the forms of carbon present. It is also shown that Raman spectroscopy can be used to follow the micromechanics of both the deformation of silicon carbide fibres and of the fibres within a model composite. Well‐defined Raman spectra have been obtained from a variety of Nicalon and Tyranno fibres and the positions of the Raman bands shown to shift on the application of stress or strain. From such stress‐induced Raman band shifts, the point‐to‐point variation of axial fibre stress or strain along an individual fibre in an epoxy matrix can be determined. An example is given of the use of the technique to map the distributions of axial fibre strain in a Nicalon/epoxy fragmentation test specimen and to model the failure processes at the fibre/matrix interface.
Microstructural analysis of Al alloys dispersed with TiB2 particulate for MMC applications
Authors: Wang  , Brydson  , Jha  , Ellis
Published: March 2003

A dispersion of TiB2 particulates in an Al alloy matrix was formed via the in‐situ reaction between mixtures of K2TiF6 (K2ZrF6), KBF4 and molten aluminium. The dispersion of the ceramic phase in the aluminium matrix was also achieved in some experiments by adding exogenous TiB2 particles to the fluoride melt in contact with molten aluminium.

In this work, we have examined the microstructure of the as‐cast metal matrix composites using analytical electron microscopy and X‐ray diffraction techniques. The phases formed as a result of the reaction between the molten fluoride flux and liquid aluminium have been identified. These were (Ti, Zr, Al)B2, Al3Ti and possibly AlB12 in the Al‐matrix, and KAlF4 and KMgF3 in the solidified flux. The mechanism of formation of TiB2 and Al3Ti is explained. The role of alloying elements is also explained in the context of interfacial chemistry and dispersion.
A simplified method of cleaning ixodid ticks for microscopy
Authors: Dixon  , Petney  , Andrews
Published: March 2003

The cleaning of ixodid ticks for microscopy can be achieved quickly and efficiently using a combination of a wax solvent and an ultrasonic cleaner. The technique involves minimum handling of specimens, produces no detectable damage and is suitable for cleaning many specimens at the same time.
Focused ion beam sample preparation of continuous fibre‐reinforced ceramic composite specimens for transmission electron microscopy
Authors: Kim  , Dravid
Published: March 2003

The microanalysis of interfaces in fibre‐reinforced composite materials is dependent on the successful preparation of specimens suitable for transmission electron microscope (TEM) inspection. Ideal samples should possess large amounts of structurally intact and uniform thin area in the fibre/matrix interface regions of the samples. Because fibre/matrix interfaces in this class of materials are often designed to fail under mechanical stress, conventionally prepared samples are prone to interfacial failure and differential thinning, both of which preclude detailed TEM microanalysis. These effects were seen in a conventionally dimpled and ion‐beam‐thinned specimen prepared from a continuous fibre reinforced ceramic composite composed of CaWO4‐coated Nextel 610TM fibres in an alumina matrix. The dimpled specimen showed large amounts of interfacial failure, with only thick regions of the specimen left intact. To overcome these limitations, a focused ion beam (FIB) technique was applied to this same material. The superiority of the FIB‐produced sample is evident in both the morphology and scanning transmission electron microscopy analyses of the sample.
Silicon technology‐based micro‐systems for atomic force microscopy/photon scanning tunnelling microscopy
Authors: Gall‐Borrut P. , Belier B. , Falgayrettes P. , Castagne M. , Bergaud C. , Temple‐Boyer P.
Published: March 2003

We developed silicon nitride cantilevers integrating a probe tip and a wave guide that is prolonged on the silicon holder with one or two guides. A micro‐system is bonded to a photodetector. The resulting hybrid system enables us to obtain simultaneously topographic and optical near‐field images. Examples of images obtained on a longitudinal cross‐section of an optical fibre are shown.
Femtosecond near‐field spectroscopy: carrier relaxation and transport in single quantum wires
Authors: Emiliani V. , Guenther T. , Lienau C. , Nötzel R. , Ploog K. H.
Published: March 2003

Quasi‐two‐colour femtosecond pump and probe spectroscopy and near‐field scanning optical microscopy are combined to study the carrier dynamics in single semiconductor nanostructures. In temporally, spectrally and spatially resolved measurements with a time resolution of 200 fs and a spatial resolution of 200 nm, the non‐linear change in reflectivity of a single quantum wire is mapped in real space and time. The experiments show that carrier relaxation into a single quantum wire occurs on a 100 fs time scale at room temperature. Evidence is given for a transient unipolar electron transport along the wire axis on a picosecond time and 100 nm length scale.
Microfabricated silicon dioxide cantilever with subwavelength aperture
Authors: Mitsuoka Y. , Niwa T. , Ichihara S. , Kato K. , Muramatsu H. , Nakajima K. , Shikida M. , Sato K.
Published: March 2003

We have developed a microfabricated SiO2 cantilever with subwavelength aperture for scanning near‐field optical microscopy (SNOM), to overcome the disadvantages of conventional optical fibre probes such as low reproducibility and low optical throughput. The microcantilever, which has a SiO2 cantilever and an aperture tip near the end of the cantilever, is fabricated in a reproducible batch process. The circular aperture with a diameter of 100–150 nm is formed by a focused ion‐beam technique. Incident light is directly focused on the aperture from the rear side of the cantilever using a focusing objective, and high optical throughput (10−2 to 10−3) is obtained. The microcantilever can be operated as a SNOM probe in contact mode or in dynamic mode.
Spectroscopic scanning near‐field optical microscopy with a free electron laser: CH2 bond imaging in diamond films
Authors: Cricenti A. , Generosi R. , Luce M. , Perfetti P. , Margaritondo G. , Talley D. , Sanghera J. S. , Aggarwal I. D. , Gilligan J. M. , Tolk N. H.
Published: March 2003

Hydrogen chemistry in thin films and biological systems is one of the most difficult experimental problems in today's science and technology. We successfully tested a novel solution, based on the spectroscopic version of scanning near‐field optical microscopy (SNOM). The tunable infrared radiation of the Vanderbilt free electron laser enabled us to reveal clearly hydrogen‐decorated grain boundaries on nominally hydrogen‐free diamond films. The images were obtained by SNOM detection of reflected 3.5 µm photons, corresponding to the C–H stretch absorption, and reached a lateral resolution of 0.2 µm, well below the λ/2 (λ= wavelength) limit of classical microscopy.
Fundamental differences between micro‐ and nano‐Raman spectroscopy
Authors: Ayars E. J. , Jahncke C. L. , Paesler M. A. , Hallen H. D.
Published: March 2003

Electric field polarization orientations and gradients close to near‐field scanning optical microscope (NSOM) probes render nano‐Raman fundamentally different from micro‐Raman spectroscopy. With x‐polarized light incident through an NSOM aperture, transmitted light has x, y and z components allowing nano‐Raman investigators to probe a variety of polarization configurations. In addition, the strong field gradients in the near‐field of a NSOM probe lead to a breakdown of the assumption of micro‐Raman spectroscopy that the field is constant over molecular dimensions. Thus, for nano‐Raman spectroscopy with an NSOM, selection rules allow for the detection of active modes with intensity dependent on the field gradient. These modes can have similar activity as infra‐red absorption modes. The mechanism can also explain the origin and intensity of some Raman modes observed in surface enhanced Raman spectroscopy.
Near‐field second harmonic imaging of the c/a/c/a polydomain structure of epitaxial PbZrxTi1–xO3 thin films
Authors: Smolyaninov I. I. , Liang H. Y. , Lee C. H. , Davis C. C. , Nagarajan V. , Ramesh R.
Published: March 2003

Near‐field optical second harmonic microscopy has been applied to imaging of the c/a/c/a polydomain structure of epitaxial PbZrxTi1–xO3 thin films in the 0 < x < 0.4 range. Comparison of the near‐field optical images and the results of atomic force microscopy and X‐ray diffraction studies show that an optical resolution of the order of 100 nm is achieved. Symmetry properties of the near‐field second harmonic signal allow us to obtain good optical contrast between the local second harmonic generation in c‐ and a‐domains. Experimentally measured near‐field second harmonic images have been compared with the results of theoretical calculations. Good agreement between theory and experiment is demonstrated.
Imaging the native structure of the chaperone protein GroEL without fixation using atomic force microscopy
Authors: Valle F. , Derose J. A. , Dietler G. , Kawe M. , Plückthun A. , Semenza G.
Published: March 2003

Most sample preparation methods for scanning probe or electron microscopy require that biomolecules, such as proteins, be fixed. Fixation destroys the molecular functionality and can possibly affect the true molecular structure. Here we report sample preparation conditions that allow the imaging of an unfixed protein, GroEL, under in‐vivo conditions, by atomic force microscopy. Under these conditions, the protein should maintain its native structure and biological activity. The typical toroidal shape with pore of the GroEL complex was easily visible in the images. Images of a single complex show dimensions that agree well with crystallographic data. Under in‐vivo conditions, it should be possible to study the biological activity and function of proteins.
Dynamic chemical mapping near a Si/SiO2 interface at elevated temperatures using plasmon‐loss images
Authors: Sasaki K. , Tsukimoto S. , Konno M. , Kamino T. , Saka H.
Published: March 2003

Plasmon‐loss imaging was applied to chemical mapping during an in‐situ heating experiment. The technique was applied to observation of vibration of a Si/SiO2 interface which took place during reduction of SiO2 at high temperature. The chemical maps of Si and SiO2 were recorded dynamically using a conventional TV‐VTR system at a time resolution of 1/30 s.
Real‐time confocal imaging, during active air abrasion – substrate cutting
Authors: Cook R. J. , Azzopardi A. , Thompson I. D. , Watson T. F.
Published: March 2003

Air abrasion cutting, using particulates accelerated in a controlled compressed gas stream, is currently being re‐evaluated as a precision tissue removal technique for dental cavity preparation. The minimal vibrations and heat generated during cutting commend the technique for use in the shaping of fragile or brittle materials that are vulnerable to vibrations and thermal stresses.

Traditional air abrasion studies have relied solely upon post‐procedure imaging, and cutting process details have been inferred from the nature of the residual surface. In this paper, however, a real‐time confocal microscopic imaging method is described, which for the first time has allowed prior target structure characterization with subsequent imaging of cutting interactions and substrate failure patterns. Using internally focusing long working distance Hill objective lenses, focusing deep to a protective microscope slide and adhesive interfaces, unhindered remote image sampling within the bulk of specimens such as tooth tissue, acrylic and brittle ceramics was possible.

Moreover, areas of active cutting and inactive regions were identified within air abraded cavities during their creation. The characteristics of the finished cut surfaces were demonstrated and confirmed the findings of previous SEM studies. The method allowed direct control over all the known variables influencing cutting with particulate streams.
Confocal laser scanning microscopy as a tool to determine cyanobacteria biomass in microbial mats
Authors: Solé A. , Gaju N. , Méndez‐Álvarez S. , Esteve I.
Published: March 2003

We have developed a method based on confocal laser scanning microscopy for detection and quantification of cyanobacteria from the Ebro Delta microbial mats. Cyanobacteria play a major role as primary producers in microbial mats; it is difficult, however, to apply classical methods to estimate their biomass because they establish strong interactions with detritic particles. The protocol described here allows the localization of individual cells or filaments with micrometre precision without the need to either manipulate or stain the samples. This method is suitable for studying biomass ‘in situ’ in microbial mats.
Electron energy‐loss near‐edge structure studies at the atomic level: reliability of the spatial difference technique
Authors: Scheu C.
Published: March 2003

The spatial difference technique was applied to determine the interface‐specific components of electron energy‐loss near‐edge structures and the results are in good agreement to those obtained by atomic column resolved measurements averaged over one atomic layer. In previous studies an experimental set‐up had been chosen where the scanning areas, which are used to measure electron energy‐loss spectra, were symmetrical in respect to the location of the investigated interfaces. In the present study, an asymmetric setting of the scanning areas was applied, which allows the interfacial signals to be determined directly. Comparing the results of the different measurements shows that the spatial difference technique is valid and can be used to obtain information about the electronic structure of interfaces where single atomic column resolved measurements are not yet possible.
Amorphous solid water produced by cryosectioning of crystalline ice at 113 K
Authors: AL‐Amoudi A. , Dubochet J. , Studer D.
Published: March 2003

Amorphous solid (vitreous) water can be obtained by a number of methods, including quick freezing of a very small volume of pure water, low pressure condensation of water vapour on a cold substrate or transformation of hexagonal ice (the ice which is naturally formed) under very high pressure at liquid nitrogen temperature. Larger volumes can be vitrified if cryoprotectant is added or when samples are frozen under high pressure. We show that a sample of 17.5% dextran solution or mouse brain tissue, respectively, frozen under high pressure (200 MPa) into cubic or hexagonal ice can be transformed into vitreous water by the very process of cryosectioning. The vitreous sections obtained by this procedure differ from cryosections obtained from vitreous samples by the irregular aspect of the sections and by small but significant differences in the electron diffraction patterns. For the growing community of cryo‐ultramicrotomists it is important to know that vitrification can occur at the knife edge. A vitreous sample is considered to show the best possible structural preservation. The sort of vitrification described here, however, can lead to bad structural preservation and is therefore considered to be a pitfall. Furthermore, we compare these sections with other forms of amorphous solid water and find it similar to high density amorphous water produced at very high pressures (about 1 GPa) from hexagonal ice and annealed close to its transformation temperature at 117 K.
The effect of the reinforcing carbon on the microstructure of pitch‐based granular composites
Authors: Méndez A. , Santamaría R. , Granda M. , Menéndez R.
Published: March 2003

Carbon composites were prepared with four pitches (a commercial impregnating coal‐tar pitch, two thermally treated pitches and an air‐blown pitch) and four granular carbons (anthracite, graphite, green petroleum coke and foundry coke). Granular carbon/pitch proportions were optimized for each composite and differed in the characteristics of the single components. Interactions of the pitch with the granular carbons during pyrolysis and their subsequent effects on the microstructure of the final composite were monitored by light microscopy. The results show that the light texture of the matrix and the porosity of the composite depend not only on the chemical composition of the pitch but also on the specific granular carbon used as reinforcing material. The same pitch may generate different light textures depending on the characteristics of the carbon. Composites from thermally treated pitches and graphite show highly ordered matrices orientated in the direction of graphite planes. Graphite particles seem to exert a huge influence on mesophase development during the pyrolysis of the treated pitches, affecting not only the orientation of the mesophase, but also reducing the rate of mesophase formation. On the other hand, when green petroleum coke is used with the thermally treated pitches, matrices show a small size light texture, due to the high reactivity of the pitch in the presence of this granular carbon. The porosity of the composites is controlled by both the pitch and the granular carbon.
Profiles of Amyloid Precursor and Presenilin 2‐Like Proteins are Correlated During Development of the Mouse Hypothalamus
Authors: Apert Caroline , Czech Christian , Faivre‐Bauman Annie , Loudes Catherine , Pradier Laurent , Epelbaum Jacques
Published: March 2003

The amyloid precursor protein (APP) and APP‐like (APLP) material, as visualized with the Mab22C11 antibody, have previously been shown to be associated with radial glia in hypothalamus, which are known to promote neurite outgrowth. By Northern blot analysis, APP 695 mRNA levels increased steadily over hypothalamic development, APP 770 mRNA was transiently expressed at 12 days postnatally, and APLP mRNA was only weakly expressed in the hypothalamus. The developmental pattern of APP moeities in mouse hypothalamus and in fetal hypothalamic neurons in culture was compared with a presenilin 2 (PS2) related protein using an antibody developed against the N‐terminal part of PS2. By Western blot analysis, APP and PS2‐like immunoreactivity were visualized as a 100–130 and 52 kDa bands, respectively. An APP biphasic increase was observed during hypothalamic development in vivo. APP immunoreactivity was equally detected in neuronal and glial cultures, while PS2‐like material was more concentrated in neurons. A correlation between APP/APP‐like and PS2‐like levels was observed during development in vivo. While APP was mostly associated with membrane fractions, a significant portion of PS2‐like material was also recovered from cytosolic fractions in vitro. In contrast to native PS2 in COS‐transfected cells, the PS2‐like material did not aggregate after heating for 90 s at 90 °C. These results indicate a close association between APP and PS2‐like material during hypothalamic development in vivo, and suggest that neuronal and glial cultures may provide appropriate models to test their interactions.
Influence of Adrenal Glands on the Modulation of Prolactin Gene Expression by the Endogenous Benzodiazepine Ligand Octadecaneuropeptide in the Male Rat Pituitary Gland
Authors: Wang Hongbo , Li Songyun , Givalois Laurent , Pelletier Georges
Published: March 2003

Recently, an 86‐amino acid polypeptide with high affinity for diazepam binding sites, termed diazepam‐binding inhibitor (DBI), has been found in the rat brain. DBI, as well as a peptide derived from DBI, the octadecaneuropeptide DBI[33–50] (ODN), interacts with the GABAA receptor complex. To investigate the role of these endogenous ligands for GABAA receptors on prolactin gene expression, we studied the effects of acute intracerebroventricular administration (4 h before sacrifice) of ODN on prolactin mRNA levels in the male rat. Because, in some neuropeptidergic systems, glucocorticoids play a role in the response to ODN, we also studied the influence of adrenal glands and the effect of dexamethasone administration in the response of prolactin gene expression to ODN. ODN injection produced an increase in prolactin mRNA levels. Adrenalectomy performed 5 days before sacrifice resulted in an increase in prolactin gene expression and also potentiated the stimulating effect of ODN. Because castration has been shown to decrease prolactin gene expression in the male rat, we used castrated and adrenalectomized animals to study the role of dexamethasone in the response of lactotrophs to ODN. In these steroid‐deprived animals, dexamethasone treatment (for 4 days) decreased prolactin mRNA levels but did not modify the response to ODN. These data indicate that an endogenous neuropeptide interacting with the GABAA receptor complex can stimulate prolactin gene expression and suggest that the adrenal glands may produce factor(s) capable of decreasing prolactin mRNA. On the other hand, it does not appear that glucocorticoid hormones play a role in the effect of ODN on lactotroph activity.
Inhibition of Naloxone‐Stimulated Adrenocorticotropin Release by Alprazolam in Myotonic Dystrophy Patients
Authors: Joyner Julie M. , Grice Jeffrey E. , Hockings Gregory I. , Torpy David J. , Crosbie Georgina V. , Walters Margaret M. , Jackson Richard V.
Published: March 2003

Myotonic dystrophy (DM) is an autosomal dominant disorder causing myotonia, progressive muscle weakness, and endocrine abnormalities including hypothalamic‐pituitary‐adrenal (HPA) axis hyperresponsiveness to CRH‐mediated stimuli. This ACTH hyperresponsiveness appears directly related to the underlying genetic abnormality. Naloxone (Nal)‐mediated CRH release causes ACTH release in normal humans and an ACTH hyperresponse in DM. Alprazolam (APZ) attenuates the ACTH release in response to Nal in normal individuals, probably by inhibiting CRH release. This study investigates the effects of APZ on Nal‐induced HPA axis stimulation in DM. The ACTH response to Nal in DM subjects was significantly reduced by APZ. Despite this DM patients have a relative resistance to APZ inhibition of Nal‐induced ACTH/cortisol release. APZ caused a smaller percentage reduction in AUC for ACTH in DM compared with controls. These findings provide further insight into the mechanism(s) of the HPA axis abnormalities in DM. In DM, there may be an increase in tonic opioid inhibition to CRH release with compensatory increases in stimulatory pathways. Alternatively, these patients may have a basal increase in pituitary vasopressin levels or an enhanced AVP/CRH synergistic mechanism at the level of the corticotroph.
Nitric Oxide Mediates Gonadotropin‐Releasing Hormne Effects on Frog Pituitary
Authors: Gobbetti Anna , Zerani Massimo
Published: March 2003

We studied the possible role of nitric oxide (NO) in GnRH‐induced gonadotropin secretion in the female water frog, Rana esculenta. During pre‐reproduction, pre‐ovulation, ovulation, post‐ovulation, refractory, recovery and hibernation, pituitaries were incubated with medium‐alone, GnRH, NO donor (NOd), NO synthase inhibitor (NOSi), cyclic GMP analogue (cGMPa), soluble guanylate cyclase inhibitor (sGCi), GnRH plus NOSi, GnRH plus sGCi, and NOd plus sGCi. Because antisera raised against gonadotropins are not available for this species, we measured these hormones indirecly through their effects on ovarian progesterone secretion. The ovaries were superfused with the pituitaries pre‐incubated as reported above. In addition, NOS activity and cGMP levels were determined in the pre‐incubated pituitaries. Those pre‐incubated with medium‐alone and with GnRH increased progesterone secretion during pre‐reproduction, pre‐ovulation, ovulation and recovery; the increase induced by GnRH was higher than that induced by medium‐alone during pre‐reproduction, pre‐ovulation and recovery. NOd and cGMPa increased progesterone in all considered reproductive phases except ovulation; the increase induced by NOd and cGMP was higher than that induced by medium‐alone during pre‐reproduction, pre‐ovulation and recovery. NOS activity was highest during ovulation and lowest during post‐ovulation, refractory and hibernation. GnRH increased NOS activity during pre‐reproduction, pre‐ovulation and recovery. Cyclic GMP levels were highest during ovulation and lowest during post‐ovulation, refractory and hibernation. GnRH increased cGMP levels during pre‐reproduction, pre‐ovulation and recovery, NOd during all considered reproductive phases. These results suggest that NO mediates basal and GnRH‐induced gonadotropin secretion in female Rana esculenta.
Role of Adrenoceptors in Vasopressin, Oxytocin and Prolactin Responses to Conditioned Fear Stimuli in the Rat
Authors: Zou  , Onaka  , Yagi
Published: March 2003

Conditioned fear or novel environmental stimuli suppress vasopressin (VP) and augment oxytocin (OT) and prolactin (PRL) release in rats. We examined the effects of intracerebroventricular (i.c.v.) injections of adrenoceptor antagonists on these neuroendocrine responses to conditioned fear or novel environmental stimuli in male rats. A β1 antagonist, metoprolol, blocked the VP but not the OT or PRL response to conditioned fear stimuli, but did not abolish neuroendocrine responses to novel environmental stimuli. A β2 antagonist, ICI118551, impaired the PRL but not the VP or OT response to fear or novel environmental stimuli. In rats injected with a α1 adrenoceptor antagonist, benoxathian, conditioned fear stimuli did not significantly induce the VP, OT or PRL responses. The effects of benoxathian were not due to a general reduction of arousal, since benoxathian did not prevent the VP, OT or PRL response to novel environmental stimuli. These data suggest that β1 adrenoceptors play a selective role in the VP response to conditioned fear stimuli, as do β2 adrenoceptors in the prolactin response to conditioned fear and novel environmental stimuli. We conclude that α1 adrenoceptors play a facilitative role in VP, OT, PRL responses to conditioned fear stimuli.
The Effects of Oestrogen and Progesterone on Serotonin and its Metabolite in the Lateral Septum, Medial Preoptic Area and Ventromedial Hypothalamic Nucleus of Female Rats
Authors: Lu  , Yuri  , Ito  , Yoshimoto  , Kawata
Published: March 2003

The effects of sex steroid hormones on serotonin and its metabolite, 5‐hydroxyindole‐3‐acetic acid (5‐HIAA) in the lateral septal nucleus (LS), the medial preoptic area (MPA) and the ventromedial nucleus of the hypothalamus (VMH) of female rats were investigated, using immunohistochemistry and high‐performance liquid chromatography (HPLC). Female rats were divided into three groups: ovariectomized rats (OVX group); OVX‐rats treated with estradiol benzoate alone (E2 group); and OVX‐rats treated with E2 plus progesterone (E2+P group). We analysed the density of serotonin‐immunoreactive fibres with a computer‐assisted image analysis system, and measured the tissue concentrations of serotonin and 5‐HIAA. Many serotonin‐immunoreactive fibres were observed in the LS, MPA and VMH in all three groups. The density of serotonin‐immunoreactive fibres in the MPA and VMH was significantly lower in the E2 and E2+P groups compared to the OVX group, whereas the LS showed no detectable differences among the three groups. In the HPLC study, the concentrations of serotonin in the MPA and VMH of the E2 and E2+P groups were significantly lower than that in the OVX group. There was no significant difference in the concentration of serotonin in the LS. The concentration of 5‐HIAA and the ratio of 5‐HIAA/serotonin in the LS, MPA and VMH showed no significant differences among the OVX, E2 and E2+P groups. The present results suggest that E2 priming for sexual behaviour can affect the serotonergic system by decreasing serotonin content, but not the turnover rate, in the MPA and VMH of female rats.
Serotonergic Innervation of the Pituitary Pars Intermedia of Xenopus Iaevis
Authors: Ubink  , Buzzi  , Cruijsen  , Tuinhof  , Verhofstad  , Jenks  , Roubos
Published: March 2003

At this point three brain centres are thought to be involved in the regulation of the melanotrope cells of the pituitary pars intermedia of Xenopus laevis: the magnocellular nucleus, the suprachiasmatic nucleus and the locus coeruleus. This study aims to investigate the existence of a fourth, serotonergic, centre controlling the melanotrope cells. In‐vitro superfusion studies show that serotonin has a dose‐dependent stimulatory effect on peptide release (1.6×basal level at 10−6 M serotonin) from single melanotrope cells. Retrograde neuronal tract tracing experiments, with the membrane probe FAST DiI applied to the pars intermedia, reveals retrogradely labelled neurones in the magnocellular nucleus, the suprachiasmatic nucleus, the locus coeruleus and the raphe nucleus. Of these brain centres, after immunocytochemistry only the raphe nucleus revealed serotonin‐immunoreactive cell bodies. In addition, serotonin‐immunoreactive cell bodies were found in the nucleus of the paraventricular organ, the posteroventral tegmental nucleus and the reticular istmic nucleus. In the pituitary, the pars nervosa, pars intermedia and pars distalis all reveal serotonin‐immunoreactive nerve fibres. With immunocytochemical double‐labelling for tyrosine hydroxylase and serotonin no colocalization of serotonin and tyrosine hydroxylase was observed in cell bodies in the brain, and in the pituitary hardly any colocalization was found in the nerve fibres. However, after in‐vitro loading of neurointermediate lobes with serotonin, tyrosine hydroxylase and serotonin appear to coexist in a fibre network in the pars intermedia. On the basis of these data we propose that the melanotrope cells in the Xenopus pars intermedia are innervated by a 5‐HT network originating in the raphe nucleus; this network represents the first identified stimulatory input to the pars intermedia of this species.
The Effects of Age and Spontaneous Adenoma Formation on Trophic Activity in the Rat Pituitary Gland: a Comparison with Trophic Activity in the Human Pituitary and in Human Pituitary Adenomas
Authors: Nolan  , Lunness  , Lightman  , Levy
Published: March 2003

The effects of ageing on trophic activity in the pituitary gland and the molecular events that underlie pituitary tumour formation are poorly understood. In the present study we have used an extremely accurate system to analyse trophic activity in human pituitary tumours and compared our findings with trophic activity in spontaneous rat pituitary adenomas and with changes in basal rates of turnover as the animals age. Thin, hematoxylin and eosin‐stained pituitary sections from groups of male Wistar rats aged 6 weeks to 16 months, killed at 90‐min intervals after receiving a single intraperitoneal bolus of colchicine to block cellular passage through mitosis, were evaluated histologically. Extremely accurate quantification of small changes in the prevalence of trophic events, and thus the rate of cell turnover, was achieved using a dedicated computerized aid to manual cell counting. Results were compared with the prevalence of mitotic activity in 24 spontaneous rat pituitary adenomas and with a series of 97 archival human pituitary adenomas and 24 normal human pituitary glands obtained at autopsy. In rats, average basal pituitary cell turnover declined by over 95% between 6 weeks and 16 months of age. Concurrent with this decline was a marked increase in the prevalence of adenoma formation. The prevalence of mitotic activity in spontaneous rat pituitary adenomas averaged almost twice that seen in normal, young rat pituitary and exceeded 16 times that seen in the pituitary of aged animals. In contrast, when compared to normal human pituitary tissue, average trophic activity in human pituitary adenomas remained extremely low.
Levels of Dopamine β Hydroxylase Immunoreactivity in the Preoptic Hypothalamus of the Ovariectomised Ewe Following Injection of Oestrogen: Evidence For Increased Noradrenaline Release Around the Time of the Oestrogen‐Induced Surge in Luteinizing Hormone
Authors: Clarke  , Scott  , Pereira  , Rawson
Published: March 2003

We have measured dopamine β hydroxylase (DBH) immunoreactivity in the preoptic hypothalamus as an index of release of noradrenaline in the ovariectomised (OVX) ewe at the time of an oestrogen‐induced surge in luteinizing hormone (LH) release. OVX ewes (n=5) were given an injection of 50 μg oestradiol benzoate (or oil), which causes a surge in the secretion of LH. Blood samples were taken and sheep were killed 16 h later. Other groups (n=3) were given oil or oestrogen and killed 6 h later. Brains were collected for immunohistochemistry and image analysis. The number of DBH‐stained cells and the integrated optical density of the cells was measured throughout the A1 field of the brainstem. The DBH staining was measured in the terminal beds of the hypothalamus. There was no difference between the controls and the EB‐treated OVX ewes in the number of DBH positive cells or the optical density of DBH‐staining cells in the A1 field. Within the preoptic area, there was reduced (P<0.02) DBH staining in the 16 h EB‐treated ewes. There was no change in the DBH staining in the paraventricular nucleus or the arcuate nucleus of the hypothalamus. These data suggest that there is release of noradrenaline in the preoptic area at the time of the E‐induced GnRH/LH surge in the OVX ewe.
Thyrotropin‐Releasing Hormone Stimulates Nitric Oxide Release from GH3 Cells
Authors: Tsumori  , Murakami  , Koshimura  , Kato
Published: March 2003

Constitutive nitric oxide synthase (NOS) is expressed in the rat adenohypophysis but the mechanisms regulating its activity at the cellular level remain to be elucidated. The effect of TRH on nitric oxide release from GH3 cells was studied by means of reverse‐phase HPLC to measure NO−2 and NO−3 concentrations in the incubation medium, and by polarography using electrodes specific for NO. Medium NO−2 concentrations in the incubation medium were dependent on the incubation time, and were further increased by sodium nitroprusside (SNP) or high potassium. NO−3 was detectable only in the presence of 100 μM SNP. Addition of
l‐arginine increased medium NO−2 concentrations. Diamino‐hydroxypyrimidine decreased medium NO−2 concentrations, which were restored by the addition of (6R)‐5, 6, 7, 8‐tetrahydro‐
l‐biopterin (THB). TRH elicited dose‐related increases in medium NO−2 concentrations and in nitric oxide‐specific currents, which were abolished by Nω‐nitro‐
l‐arginine methyl ester. TRH failed to increase medium NO−2 concentrations in cells loaded with an intracellular Ca2+‐chelating agent. The findings suggest that mobilization of intracellular Ca2+ by TRH stimulation activates Ca2+‐dependent NOS in GH3 cells.
The Progesterone Blockade of the Luteinizing Hormone Surge is Overcome by RU486
Authors: Skinner  , Bouchard  , Caraty
Published: March 2003

Progesterone can prevent the oestrogen‐induced and spontaneous preovulatory luteinizing hormone (LH) surges but the mechanisms underlying this effect remain poorly understood. Using a follicular phase ovariectomised (OVX) ewe model and by elevating progesterone in the presence of oestrogen to inhibit the LH surge, we investigated whether the progesterone receptor antagonist, RU486, could block the inhibitory effects of progesterone. Accordingly, intravaginal progesterone implants were inserted into OVX Ile‐de‐France ewes (n=18), bearing 10 mm Silastic 17β‐oestradiol implants. Ten days later, the progesterone implants were removed, whereupon new implants were inserted immediately into 12 ewes: six of which were also injected with 100 mg RU486 dissolved in 10 mL vehicle (10% alcohol in peanut oil) and six received vehicle only. The remaining six ewes were injected with vehicle only. RU486 and vehicle injections were made again 12 and 24 h later. After the last injection, oestrogen concentrations were raised to peak follicular phase levels in all ewes by subcutaneous insertion of four 3‐cm 17β‐oestradiol implants. Blood samples were collected every 2 h for 40 h starting 9 h after the insertion of the oestrogen implants. As expected, the six ewes treated only with oestradiol had a LH surge whereas no ewes given the implants in the presence of progesterone surged. RU486 completely blocked the inhibitory effect of progesterone. There were no differences in the time of LH surge onset, duration over which LH levels remained above their half‐maximal concentration or magnitude of the LH surge between the two groups showing surges. Our study suggests strongly that the progesterone‐mediated blockade of the ovine oestrogen‐induced LH surge is not through allopregnanolone activation of the GABAA receptor. Rather, our study demonstrates that this effect is transduced by the classic nuclear progesterone receptor.
The Effect of GHRP‐6 on the Intracellular Na+ Concentration of Rat Pituitary Cells in Primary Culture
Authors: Kato  , Sakuma
Published: March 2003

The objective of the present study was to further investigate the ionic mechanism of the action of GHRP‐6 on male rat pituitary cells in culture. A synthetic hexapeptide, GHRP‐6 stimulates the secretion of growth hormone both in vivo and in vitro. It is generally accepted that Ca2+ and protein kinase C but not cAMP are involved in the signal transduction pathway of the action of GHRP‐6. Ca2+‐influx through voltage‐gated Ca2+ channels and mobilization of internal stored Ca2+ are thought to be responsible for an increase in cytosolic Ca2+ concentration. For activation of the voltage‐gated Ca2+ channels, however, it is not determined whether the membrane Na+ permeability plays a role. To answer this question, we measured intracellular Na+ concentration of the pituitary cells with ion imaging technique. We found that GHRP‐6 increased [Na+]i; the Na+ response depended on the presence of extracellular Na+ and was blocked by Gd3+, known as a blocker of nonselective cation channels but not by tetrodotoxin, a blocker of the voltage‐gated Na+ channel; thapsigargin, an inhibitor of endoplasmic reticulum Ca2+ ATPase, had no effect on the response; Ca2+ chelating agent, BAPTA had no inhibitory effect on the response; ouabain, an inhibitor of Na+‐K+ ATPase, did not block the rise in [Na+]i induced by GHRP‐6; somatostatin, which hyperpolarizes the cells by activating K+ channels, suppressed the response. These data clearly showed that GHRP‐6 increased [Na+]i in the rat pituitary cells including somatotrophs. The rise in [Na+]i is likely to be due to an increase in the membrane Na+ permeability which should depolarize the cells, thereby activating the voltage‐gated Ca2+ channels. This process leads to an influx of Ca2+ and subsequent increase in [Ca2+]i which results in an exocytotic release of GH.
Long‐Term Treatment with Antidepressants Increases Glucocorticoid Receptor Binding and Gene Expression in Cultured Rat Hippocampal Neurones
Authors: Okugawa  , Omori  , Suzukawa  , Fujiseki  , Kinoshita Toshihiko , Inagaki Chiyoko
Published: March 2003

Since the glucocorticoid receptor (GR) and/or mineralocorticoid receptor (MR) in the hippocampus have been implicated in cortisol feedback of the hypothalamus‐pituitary‐adrenal (HPA) axis, abnormalities in those receptors might underlie the hyperactivity of the HPA axis described in patients with major depression. Animal studies have shown that long‐term in‐vivo treatment with antidepressants up‐regulates hippocampal GR and/or MR, but it is not clear whether this up‐regulation is evoked through a direct action of antidepressants on these receptors. We therefore examined the direct effects of long‐term antidepressant treatment on GR binding and the levels of GR messenger RNA (mRNA) in primary cultures of rat hippocampal neurones. The time course of the effects of the tricyclic antidepressants desipramine and amitriptyline on GR binding, as assessed by [3H]dexamethasone binding using RU 28362, a specific agonist for GR, showed a biphasic mode of stimulation: desipramine significantly increased the GR binding with 2‐day exposure by 36% over that in controls and by 99% and 60% with 10‐ and 14‐day exposures, respectively. Amitriptyline also led to a significant increase in GR binding, with peaks at 2 (by 60%) and 14 days of exposure (by 60%). The effects of 14‐day treatment with desipramine required at least the first 4‐day exposure, and the first 10‐day exposure was required for the full effect. Northern blot analysis demonstrated that the GR mRNA level was significantly increased by 14‐day treatment with desipramine (+142% over control), amitriptyline (+108%), mianserin (+124%), paroxetine (+42%) and sulpiride (+92%), but not with haloperidol. Immunocytochemistry for GR revealed that 2‐ or 14‐day treatment with desipramine significantly increased the number of GR‐positive cells with dominant immunoreactivity in the nuclei of granule cell‐like neurones or in perikarya of pyramidal cell‐ and granule cell‐like neurones. These findings suggest that tricyclic antidepressants directly increase hippocampal GR by short‐term (2‐day) and long‐term (14‐day) exposure, and that the increase by long‐term exposure is evoked commonly with different classes of antidepressants through transcriptional up‐regulation of GR expression.
Stimulatory Role of Prolactin on the Development of Tuberoinfundibular Dopaminergic Neurones in Prepubertal Female Rats: Studies with Cysteamine and Somatostatin
Authors: Shieh  , Pan
Published: March 2003

Cysteamine, a potent depletor of prolactin and somatostatin, was used to determine the role of prolactin and somatostatin in the control of central dopamine neurones in prepubertal rats. Cysteamine (100 mg/kg, i.p., twice daily) was injected for 7, 14 or 21 days in 28‐day‐old Sprague‐Dawley female rats in one study and for 3 days in 35‐day‐old rats in another. In control rats, the 3,4‐dihydroxyphenylacetic acid (DOPAC) levels in the median eminence increased threefold from day 35 to day 49, and serum prolactin concentration increased about 50%. Cysteamine lowered serum prolactin concentrations to 20%, and median eminence DOPAC and dopamine levels to 32–50% of control levels in both studies. The DOPAC levels in the nucleus accumbens and striatum were also lowered, while both DOPAC and dopamine in the paraventricular nucleus and periventricular nucleus (A14) were increased by cysteamine. A single injection of rat prolactin (0.01, 0.1 or 1 mg/kg) significantly increased DOPAC or DOPA levels in the median eminence, nucleus accumbens and striatum, but not in the paraventricular nucleus or A14 at 14 h later in 28‐day old female rats or in 40‐day‐old rats pretreated with cysteamine. In contrast, central injection of somatostatin dose (0.001–1 μg/rat) and time (30–90 min) dependently decreased the DOPAC levels in the median eminence, paraventricular nucleus and A14 and increased those in the nucleus accumbens and striatum of adult female rats. These results indicate that serum prolactin is important for the maturation and maintenance of dopamine systems in the median eminence, nucleus accumbens and striatum, while somatostatin exhibits inhibitory and stimulatory effects on hypothalamic and midbrain dopamine systems, respectively.