bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of the ERK homologue CpMK2 from the chestnut blight fungus Cryphonectria parasitica
Authors: Choi Eun-Sil , Chung Hea-Jong , Kim Myoung-Ju , Park Seung-Moon , Cha Byeong-Jin , Yang Moon-Sik , Kim Dae-Hyuk
Published: May 2005

The Cryphonectria parasitica gene cpmk2, which encodes a mitogen-activated protein kinase belonging to the yeast extracellular signalling-regulated kinase (YERK1) subfamily, was isolated and its biological function was examined. Disruption of cpmk2 resulted in impaired pigmentation and abolished conidiation. Growth defects were observed in the cpmk2 mutant grown on solid plates, but growth of the mutant appeared normal in liquid media, including EP complete and PD broth, suggesting that the cpmk2 gene is involved in sensing and responding to growth conditions. The mutant's production of laccase, as measured by the size of the coloured area produced on tannic-acid-supplemented plates, was significantly reduced compared with the wild-type, but the intensity of the coloured area was unchanged, suggesting that the reduced laccase activity was owing to reduced growth on solid media rather than transcriptional downregulation. A dramatic reduction observed in the canker area produced by the cpmk2 mutant compared with the wild-type, even more severe than that of a hypovirulent strain, can also be ascribed to defective growth on solid surfaces rather than to impairments in a virulence factor(s). Downregulation of the pheromone gene Mf2/1 was also observed in the mutant, indicating a possible explanation for the regulation of the pheromone precursor gene in filamentous fungi and suggesting the presence of the yeast-like pheromone-responsive pathway in C. parasitica. Immunoblot analyses revealed that the phosphorylation level of CpMK2 increased in both virus-free and virus-containing strains in liquid cultures of up to 5 days old and decreased in older cultures. Moreover, the CpMK2 phosphorylation level increased in both strains after transfer from liquid to solid medium. However, levels of phosphorylated CpMK2 were similar in the two strains, suggesting that CpMK2, unlike CpMK1, is not under the direct control of a hypovirus.
Ethambutol, a cell wall inhibitor of Mycobacterium tuberculosis, elicits l-glutamate efflux of Corynebacterium glutamicum
Authors: Radmacher Eva , Stansen Kathrin C. , Besra Gurdyal S. , Alderwick Luke J. , Maughan William N. , Hollweg Günter , Sahm Hermann , Wendisch Volker F. , Eggeling Lothar
Published: May 2005

Corynebacterium glutamicum is used for the large-scale production of l-glutamate, but the efflux of this amino acid is poorly understood. This study shows that addition of ethambutol (EMB) to growing cultures of C. glutamicum causes l-glutamate efflux at rates of up to 15 nmol min−1 (mg dry wt)−1, whereas in the absence of EMB, no efflux occurs. EMB is used for the treatment of Mycobacterium tuberculosis, and at a molecular level it targets a series of arabinosyltransferases (EmbCAB). The single arabinosyltransferase-encoding emb gene of C. glutamicum was placed under the control of a Tet repressor (TetR). Experiments with this strain, as well as with an emb-overexpressing strain, coupled with biochemical analyses showed that: (i) emb expression was correlated with l-glutamate efflux, (ii) emb overexpression increased EMB resistance, (iii) EMB caused less arabinan deposition in cell wall arabinogalactan, and (iv) EMB caused a reduced content of cell-wall-bound mycolic acids. Thus EMB addition resulted in a marked disordering of the cell envelope, which was also discernible by examining cellular morphology. In order to further characterize the cellular response to EMB addition, genome-wide expression profiling was performed using DNA microarrays. This identified 76 differentially expressed genes, with 18 of them upregulated more than eightfold. Among these were the cell-wall-related genes ftsE and mepA (encoding a secreted metalloprotease); however, genes of central metabolism were largely absent. Given that an altered lipid composition of the plasma membrane of C. glutamicum can result in l-glutamate efflux, we speculate that major structural alterations of the cell envelope are transmitted to the membrane, which in turn activates an export system, perhaps via increased membrane tension.
Biochemical and molecular characterization of an azoreductase from Staphylococcus aureus, a tetrameric NADPH-dependent flavoprotein
Authors: Chen Huizhong , Hopper Sherryll L. , Cerniglia Carl E.
Published: May 2005

Azo dyes are a predominant class of colourants used in tattooing, cosmetics, foods and consumer products. A gene encoding NADPH-flavin azoreductase (Azo1) from the skin bacterium Staphylococcus aureus ATCC 25923 was identified and overexpressed in Escherichia coli. RT-PCR results demonstrated that the azo1 gene was constitutively expressed at the mRNA level in S. aureus. Azo1 was found to be a tetramer with a native molecular mass of 85 kDa containing four non-covalently bound FMN. Azo1 requires NADPH, but not NADH, as an electron donor for its activity. The enzyme was resolved to dimeric apoprotein by removing the flavin prosthetic groups using hydrophobic-interaction chromatography. The dimeric apoprotein was reconstituted on-column and in free stage with FMN, resulting in the formation of a fully functional native-like tetrameric enzyme. The enzyme cleaved the model azo dye 2-[4-(dimethylamino)phenylazo]benzoic acid (Methyl Red) into N,N-dimethyl-p-phenylenediamine and 2-aminobenzoic acid. The apparent K
m values for NADPH and Methyl Red substrates were 0·074 and 0·057 mM, respectively. The apparent V
max was 0·4 μM min−1 (mg protein)−1. Azo1 was also able to metabolize Orange II, Amaranth, Ponceau BS and Ponceau S azo dyes. Azo1 represents the first azoreductase to be identified and characterized from human skin microflora.
Genetic analysis of Bacillus anthracis Sap S-layer protein crystallization domain
Authors: Candela Thomas , Mignot Tâm , Hagnerelle Xavier , Haustant Michel , Fouet Agnès
Published: May 2005

Bacillus anthracis, the aetiological agent of anthrax, synthesizes two surface-layer (S-layer) proteins. S-layers are two-dimensional crystalline arrays that completely cover bacteria. In rich medium, the B. anthracis S-layer consists of Sap during the exponential growth phase. Sap is a modular protein composed of an SLH (S-layer homology)-anchoring domain followed by a putative crystallization domain (Sapc). A projection map of the two-dimensional Sap array has been established on deflated bacteria. In this work, the authors used two approaches to investigate whether Sapc is the crystallization domain. The purified Sapc polypeptide (604 aa) was sufficient to form a crystalline structure, as illustrated by electron microscopy. Consistent with this result, the entire Sapc domain promoted auto-interaction in a bacterial two-hybrid screen developed for the present study. The screen was derived from a system that takes advantage of the Bordetella pertussis cyclase subdomain structure to enable one to identify peptides that interact. A screening strategy was then employed to study Sapc subdomains that mediate interaction. A random library, derived from the Sapc domain, was constructed and screened. The selected polypeptides interacting with the complete Sapc were all larger (155 aa and above) than the mean size of the randomly cloned peptides (approx. 60 residues). This result suggests that, in contrast with observations for other interactions studied with this two-hybrid system, large fragments were required to ensure efficient interaction. It was noteworthy that only one polypeptide, which spanned aa 148–358, was able to interact with less than the complete Sapc, in fact, with itself.
Distinctive Processing Underlies Skilled Memory
Authors: Van Overschelde James P. , Rawson Katherine A. , Dunlosky John , Hunt R. Reed
Published: May 2005

Research on skilled memory has focused on organizational processes to the exclusion of item-specific processes, although theories of skilled memory do acknowledge the importance of both kinds of processes. Using the isolation methodology, we presented lists of American football team names to participants who had either a high or a low level of knowledge about American football. An isolation effect (greater recall of the target in the isolate list than in a homogeneous control list) was observed only with high-knowledge participants. When standard lists were used, an isolation effect was observed with both groups. These findings empirically validate the importance of both organizational and item-specific processing as the basis of distinctive processing underlying skilled memory performance.
Do Speakers Avoid Ambiguities During Dialogue?
Authors: Haywood Sarah L. , Pickering Martin J. , Branigan Holly P.
Published: May 2005

What affects speakers' production of ambiguous utterances in dialogue? They might consider ease of production for themselves, or ease of comprehension for their addressees. Previous research has demonstrated that ease of production plays a role in speakers' syntactic choices, but that ease of comprehension does not. However, such studies have not employed dialogues that involve role swapping on a turn-by-turn basis. In our experiment, participants alternated in giving and following instructions to move objects around on a grid. They tended to repeat the syntactic form just used by their interlocutor, reflecting sensitivity to ease of production. More interestingly, they were more likely to disambiguate their utterances when the visual context was potentially ambiguous than when it was not, reflecting sensitivity to ease of comprehension. We conclude that speakers pay attention to their beliefs about their addressees' ease of comprehension, in addition to considering ease of production for themselves.
Effects of size and fragmentation of marine reserves and fisher infringement on the catch and biomass of coral trout, Plectropomus leopardus, on the Great Barrier Reef, Australia
Authors: LITTLE L. R. , SMITH A. D. M. , MCDONALD A. D. , PUNT A. E. , MAPSTONE B. D. , PANTUS F. , DAVIES C. R.
Published: May 2005

Abstract  A spatially structured simulation model of the population dynamics and line fishing exploitation of common coral trout, Plectropomus leopardus Lacepède, was used to evaluate the effects of infringement and different amounts and arrangements of marine reserves on the Reef Line Fishery of the Great Barrier Reef, Australia. With no marine reserves and under a constant future effort level equal to that for 1996, the size of the population was reduced and the biomass stabilised at about 40% of pre‐exploitation levels. Marine reserves were ineffective at conserving biomass when limited infringement was allowed throughout an entire reserve. When infringement was absent altogether or limited to the edges of reserves, larger marine reserves lead to lower total catches and higher overall biomass. When infringement was limited to reserve edges, a single large closure was more effective at conserving biomass than more fragmented arrangements. Simulations suggested that marine reserves might lead to better conservation of a fishery‐targeted species if infringement is negligible or limited to reserve margins. Even where infringement occurred only at the edges of reserves, a network of small reserves may be less effective at conserving a targeted species than a smaller number of larger reserves.
Regulation of Peroxisome Proliferator Activated Receptor‐γ in Rat Pituitary
Authors: Nogueiras R. , Caminos J. E. , Gallego R. , Raghay K. , Bravo S. , Tovar S. , Pombo C. , López M. , Tena‐Sempere M. , García‐Caballero T. , Diéguez C.
Published: May 2005

Peroxisome proliferator activated‐receptor γ (PPARγ) is a member of the nuclear receptor superfamily and, in addition to its relation with obesity and insulin sensitivity, it has recently been localized in human and mice pituitary, indicating a functional significance of PPARγ in adenopituitary tumours. In the present study, we localized the PPARγ mRNA and protein in different cell types of rat pituitary. Moreover, using the real‐time polymerase chain reaction, we assessed the mRNA expression of PPARγ in different physiological and pathological settings known to be associated with alterations in anterior pituitary cell proliferation and/or function. Our experiments have shown that PPARγ mRNA levels were repressed by oestrogen through an oestrogen receptor‐α effect. However, PPARγ protein levels were only modified in males but not in females. On the other hand, PPARγ mRNA expression was increased in dwarf rats in comparison with Lewis rats. Finally, nutritional, thyroid status or pregnancy did not change PPARγ expression. Taken together, we provide new data regarding the regulation of pituitary PPARγ mRNA by hormonal and metabolic status.
Effect of Prolyl Endopeptidase Inhibition on Arginine‐Vasopressin and Thyrotrophin‐Releasing Hormone Catabolism in the Rat Brain
Authors: Bellemère G. , Vaudry H. , Morain P. , Jégou S.
Published: May 2005

Compound S 17092 is a potent and selective inhibitor of prolyl endopeptidase (EC 3.4.21.26, PEP) that may be of therapeutic value for the treatment of memory impairment associated with neurodegenerative diseases. In the present study, we investigated the effects of S 17092 on the catabolism of the promnesic neuropeptides thyrotrophin‐releasing hormone (TRH) and arginine‐vasopressin (AVP) in the rat brain. In vitro, bacterial PEP hydrolysed both TRH and AVP, and the breakdown of the two peptides was almost completely prevented by 10−5 M S 17092. In vivo, a single oral administration of S 17092 provoked a significant increase in TRH‐like immunoreactivity (TRH‐LI) in the cerebral cortex (+63% for a 10 mg/kg dose and +72% for a 30 mg/kg dose), as well as AVP‐LI in the hippocampus (+54% for a 30 mg/kg dose), but did not affect TRH‐LI in the amygdala nor AVP‐LI in the cerebral cortex. Chronic administration of S 17092 (10 or 30 mg/kg daily) lead to a significant increase in THR‐LI in the cerebral cortex (+55% and +56%, respectively), but did not modify AVP‐LI in the hippocampus, nor in the cerebral cortex. These results show that the selective PEP inhibitor S 17092 increases TRH and AVP content in discrete regions of the rat brain. The present data suggest that the promnesic and antiamnesic effects of S 17092 can be accounted for, at least in part, by blockage of AVP and TRH degradation by PEP.
Expression of Functional Ciliary Neurotrophic Factor Receptors in Immortalized Gonadotrophin‐Releasing Hormone‐Secreting Neurones
Authors: Dozio E. , Watanobe H. , Ruscica M. , Maggi R. , Motta M. , Magni P.
Published: May 2005

Ciliary neurotrophic factor (CNTF), a cytokine of the interleukin‐6 superfamily, is known to exert pleiotropic actions, including regulation of food intake and permissive effects on reproduction, by facilitating the release of gonadotrophin‐releasing hormone (GnRH) and gonadotrophins. CNTF activates membrane receptors (CNTF‐Rs) composed of one ligand‐specific binding subunit, defined CNTFRα, and two signal transducing subunits, termed leukaemia inhibitory factor receptor (LIFR) and gp130. However, it is not clear whether the effects of CNTF on GnRH release result from either a direct or an indirect action on GnRH‐secreting hypothalamic neurones, or from a combination of these events. The hypothesis of a direct effect of CNTF was thus tested using the GT1‐7 GnRH‐secreting cell line. CNTF‐R expression and CNTF‐induced modulation of the Janus kinase (JAK2)‐signal transducer and activator of transcription 3 (STAT3) pathway and of GnRH release were evaluated. GT1‐7 cells were found to express CNTFRα, LIFR and gp130 genes, as shown by reverse transcription‐polymerase chain reaction analysis, and the corresponding proteins, analysed by immunofluorescence and western blot. CNTFRα, LIFR and gp130 immunoreactive bands had an approximate size of 50, 190 and 130 kDa, respectively. Treatment of GT1‐7 cells with 10−12 M CNTF for 15–60 min resulted in a marked and transient increase of STAT3 phosphorylation via activation of JAK2. A 30‐min exposure of GT1‐7 cells to different CNTF concentrations increased the accumulation of GnRH into the culture medium, with a maximal effect at 10−11 M. In conclusion, the present results provide new information about the regulation of the reproductive axis by CNTF, and suggest that it might operate at the hypothalamic level by directly influencing the activity of GnRH‐secreting neurones, in addition to the possible indirect effects via interneurones proposed by previous studies.
GABAergic Modulation of the Expression of Genes Involved in GABA Synaptic Transmission and Stress in the Hypothalamus and Telencephalon of the Female Goldfish (Carassius auratus)
Authors: Martyniuk C. J. , Crawford A. B. , Hogan N. S. , Trudeau V. L.
Published: May 2005

GABA is one of the most abundant neurotransmitters in the vertebrate central nervous system and is involved in neuroendocrine processes such as development, reproduction, feeding and stress. To examine the effect of GABA on gene expression in the brain, we used a cDNA macroarray containing 26 genes involved in GABA synaptic transmission (GABA receptor subunits, GABA transporters), reproduction (gonadotrophin‐releasing hormone isoforms and oestrogen receptor α), feeding (neuropeptide Y and cholecystokinin), and stress [corticotrophin‐releasing factor (CRF)]. To elevate GABA levels in the brain, we injected female goldfish with gamma‐vinyl GABA (300 µg/g of body weight) (24 h), an irreversible inhibitor of the enzyme GABA transaminase (GABA‐T). We found that increased levels of GABA in the hypothalamus resulted in a 2.2‐fold down‐regulation of GABAA receptor β4 subunit mRNA. In the telencephalon, we found that increased GABA levels resulted in a 1.5‐fold increase of CRF mRNA and a 1.8‐fold decrease of GABAA receptor β2 subunit mRNA. Increasing GABA in the hypothalamus and telencephalon of the goldfish did not significantly affect the mRNA abundance of genes involved in GABA synthesis (glutamic acid decarboxylase isoforms) and degradation (GABA‐T), feeding, or reproduction. Our preliminary study suggests that the regulation of GABA receptor subunit mRNA expression by GABA may be a conserved evolutionary mechanism in vertebrates to modulate GABAergic synaptic transmission.
Subduction of arc basaltic andesite: implications for the tectonic history of the southern New England Fold Belt
Authors: OFFLER R. , GLEN R. A. , HYODO H. , JIANG Z.
Published: May 2005

Serpentinite mélange in the Peel–Manning Fault System, southern New England Fold Belt hosts tectonic blocks of varying size and lithology. One of the largest occurs at Woodsreef, northern New South Wales, as multiply deformed amphibole schists recording at least three phases of deformation (D1–D3). They have a peralkaline, composition (molecular Na2O+K2O = 0.083–0.189; Al2O3 = 0.046–0.130) and contain aegirine, quartz, albite, arfvedsonite and less common actinolite, neptunite, aenigmatite and bafertisite, as well as relict pre‐D1 aegirine. The Zr/TiO2and Nb/Y ratios suggest that the protoliths had originally basaltic to basaltic andesite composition, indicating that they have been altered to a rock of peralkaline composition. Chondrite normalised patterns [(Ce/Yb)N = 4.98–9.36], Hf/3–Th–Ta, Hf/3–Th–Nb/16 and Y/15–La/10–Nb/8 discrimination diagrams, and rock/MORB plots indicate that the protoliths had a calc‐alkaline, arc composition.143Nd/144Nd data (εNd(500) = 2.8–3.3; −4.8; εNd(400) = 2.0–2.5; −5.8) imply that they have been derived from isotopically mildly depleted and enriched sources. Single‐grain40Ar–39Ar dating of felsic and mafic fractions from one sample, produces ages of 1200–1100 Ma (albite–quartz vein) and 335.2 ± 2.1 Ma, 332 ± 3.7 Ma and 291 ± 3.7 Ma (arfvedsonite‐rich matrix). The Neoproterozoic age obtained from the felsic fraction is attributed to the presence of excess40Ar in quartz and albite, and the 291 Ma age to partial resetting. The 335–332 Ma ages are interpreted as cooling ages related to uplift after subduction. The basalts and basaltic andesites underwent alteration either within a volcanic pile or during subduction after collision with a ?west‐dipping subduction zone at the proto‐Gondwana margin. With continuing subduction, they were recrystallised and deformed in the deeper levels of the subduction zone, producing aegirine with up to 38% jadeite in solid solution. Replacement of the high‐P assemblages by lower P/high‐T arfvedsonite‐bearing assemblages then took place during uplift atca 334 Ma.
Carbonate sediments of Elizabeth and Middleton Reefs close to the southern limits of reef growth in the southwest Pacific
Authors: KENNEDY D. M. , WOODROFFE C. D.
Published: May 2005

Elizabeth and Middleton Reefs are atoll‐like structures that have developed on top of volcanic edifices and are close to the southern environmental limit of reef development in the southwest Pacific. Reef morphology and vertical accretion rates during the Holocene appear similar to those on other more tropical reefs. Sediment samples were collected from the lagoon of both reefs and around the flanks of Middleton Reef. A distinctly chlorozoan assemblage was observed with coral, molluscs, Halimeda, coralline algae and foraminifers being the dominant sediment constituents. Lagoon sediment samples show little variation within or between reefs, lacking the concentric zonation characteristic of larger atolls. Samples collected from the flanks of Middleton Reef, and subsurface material from vibrocores, differ compositionally from the surficial lagoon sand and were typically more tropical in character. A comparison of the sediment constituents from these reefs with those of samples from within a fringing reef and from the shelf around Lord Howe Island, further south, indicated regional patterns in sediment composition. Halimeda rapidly decreased in abundance with increased latitude, and appeared confined to deeper water, whereas coralline red algae increased significantly. The rapid change in these major sediment contributors is coincident with the general decrease in coral growth rates with latitude. This reinforces the notion that the latitudinal limit of reef development is constrained by factors other than coral growth alone.
The Narooma Terrane: implications for the construction of the outboard part of the Lachlan Orogen
Authors: GLEN R. A. , STEWART I. R. , PERCIVAL I. G.
Published: May 2005

In its type area around Narooma, the Narooma Terrane in the Lachlan Orogen comprises the Wagonga Group, which consists of the Narooma Chert overlain by the argillaceous Bogolo Formation. Conodonts indicate that the lower, largely massive (ribbon chert) part of the Narooma Chert ranges in age from mid‐Late Cambrian to Darriwilian–Gisbornian (late Middle to early Late Ordovician). The upper Narooma Chert consists of shale, containing Eastonian (Late Ordovician) graptolites, interbedded with chert. Where not deformed by later faulting, the boundary between the Narooma Chert and Bogolo Formation is gradational. At map scale, the Narooma Terrane consists of a stack of imbricate thrust slices caught between two thrust faults that juxtaposed the terrane against the coeval Adaminaby Superterrane in Early Silurian time. These slices are best defined where Narooma Chert is thrust over Bogolo Formation. The soles of such slices contain multiply foliated chert. Late extensional shear bands indicate a strike‐slip component to the faulting. The Narooma Terrane, with chert overlain by muddy ooze, is interpreted to be an oceanic terrane that accumulated remote from land for ∼50 million years. The upward increase in the terrigenous component at the top of the Wagonga Group (shale, argillite, siltstone and sandstone of the upper Narooma Chert and Bogolo Formation) records approach of the terrane to the Australian sector of the Gondwana margin. Blocks of chert, argillite and sandstone reflect extensional/strike‐slip disruption of the terrane as it approached the transform trench along the Gondwana–proto‐Pacific plate boundary. Blocks of basalt and basalt breccia represent detritus from a seamount that was also entering the trench. There is no evidence that the Narooma Terrane or the adjacent Adaminaby Group formed in an accretionary prism/subduction complex.
Changes in the heart rate and plasma epinephrine and norepinephrine concentrations of the stallion during copulation
Authors: TERADA MISAO , MOMOZAWA YUKIHIDE , KOMANO MICHIO , KUSUNOSE RYO , SATO FUMIO , SAITO TORU R.
Published: May 2005

Aim: The physiological indexes of the copulatory behavior of the stallion have not been investigated in detail and may differ from those of other species, such as humans and rats.

Methods: In order to understand the breeding capability of various stallions, their behavior during copulation was observed, and heart rate (HR) and the plasma concentrations of norepinephrine (NA) and epinephrine (Ad) were measured sequentially for a total of 13 copulations carried out during 2 days.

Results: The mean HR at rest was 35.3 ± 0.9 beats per minute (b.p.m.) and it peaked during mounting (162.1 ± 5.4 b.p.m.). The HR at ejaculation was 145.7 ± 5.1 b.p.m, which was less than the peak. The plasma concentrations of NA and Ad showed similar changes to the HR; immediately after ejaculation they were, respectively, 4.7‐fold and 1.9‐fold higher than the resting values and there was a difference in the degree of increase of each catecholamine.

Conclusions: The present results show that in the stallion the HR peaks at mounting and there is a greater change in the activity of the sympathetic nervous system because of the short‐term, highly intense exercise performed during copulation. It is considered that this, combined with the particular mental stress placed on the stallion during copulation, has the potential to cause sudden cardiac death. (Reprod Med Biol 2005;4: 143–148)
Pregnancy rate, multiple pregnancy rate, and embryo quality: Clues for single blastocyst transfer from double blastocyst transfer in an unselected population
Authors: HARA TETSUAKI , KATSUKI TAKAFUMI , KUSUDA TOMOYO , OHAMA KOSO
Published: May 2005

Objective:  Minimizing multiple pregnancy is a priority in assisted reproduction. As implantation rates are critical to success and reduce multiple pregnancy, we investigated whether blastocyst grade determined implantation rate following double blastocyst transfer in unselected cases.

Materials and Methods:  We studied 69 three‐cleavage stage embryo transfers and 64 two‐blastocyst transfers. Two blastocysts, or one when two blastocysts were not available, were transfered after evaluating the grade of blastocysts. The difference in pregnancy and implantation rates to patient age, the number of retrieved oocytes and grade of blastocysts were analyzed.

Results:  Blastocyst and grade 3AA rates per fertilized egg were 50.3% and 26.0%, respectively. Following two‐blastocyst transfer, pregnancy rate per transfer, implantation rate per embryo, and multiple pregnancy rate per pregnancy were 39.1%, 26.5%, and 24.0%, respectively. Two‐blastocyst transfer achieved implantation more often than three‐cleavage‐stage embryo transfer, but did not reduce multiple pregnancy. Pregnancy, implantation, and multiple pregnancy rates did not reflect maternal age. Higher pregnancy and implantation rates per transfer were attained for with six or more oocytes retrieved or transfer of two‐blastocyst graded 3AA or higher especially when two or more blastocysts graded 3AA or higher are available, but the latter showed a high multiple pregnancy rate (38.5%).

Conclusions:  Single embryo transfer could be carried out when two or more blastocysts of grade 3AA or higher have been developed. (Reprod Med Biol 2005; 4: 153–160)
Survival of mouse ovarian tissue transplanted into the uterine horn of post‐partum rats nursing pups of various numbers and sizes
Authors: KAGABU SATOSI , UMEZU MOTOAKI
Published: May 2005

Aim and background:  To investigate the effect of the number of pups being nursed on the survival of mouse ovarian tissue transplanted into the post‐partum rat uterus.

Methods:  Mouse ovarian tissue was transplanted into the uterine horn of post‐partum rats. The number of pups nursed by each recipient rat was adjusted in a manner predetermined. Examinations were undertaken at 1–11 weeks after transplantation. Ovarian tissue containing healthy follicles was considered to have taken successfully.

Results:  In rats with 12 pups, ovarian tissue remained viable at 11 weeks post‐transplantation. No viable ovarian tissue remained when there were one or two pups. Viability improved as the number of pups increased.

Conclusion:  When mouse ovarian tissue is transplanted into post‐partum lactating rats, viability improves as the number of pups increases. We concluded that these findings may be explained in terms of progesterone levels in the recipient rats. (Reprod Med Biol 2005; 4: 149–152)
Microtubule organization during rabbit fertilization by intracytoplasmic sperm injection with and without sperm centrosome
Authors: MORITA JUNKO , TERADA YUKIHIRO , HOSOI YOSHIHIKO , FUJINAMI NAHOKO , SUGIMOTO MIKI , NAKAMURA SOU‐ICHI , MURAKAMI TAKASHI , YAEGASHI NOBUO , OKAMURA KUNIHIRO
Published: May 2005

Aim: In most mammalian fertilization, the sperm introduces the centrosome, which acts as a microtubule organizing center (MTOC) and is essential for pronuclear movement. In rabbit fertilization, biparental centrosomal contribution in microtubule organization has been suggested.

Methods: To reveal the function and inheritance of the centrosome during rabbit fertilization, we compared microtubule organization and early embryonal development following intracytoplasmic sperm injection (ICSI) with and without sperm centrosome. Sperm centrosome was removed by sonication, and the isolated sperm head was injected by a Piezo‐driven micromanipulator. Samples were studied by light microscope after immunocytological stain.

Results: The sperm aster formation was observed 2–3 h after ICSI with intact sperm. In contrast, microtubules were organized between the male and female pronucleus without a nucleation site in the eggs after ICSI with an isolated sperm head. In the late pronuclear stage following ICSI with an isolated sperm head, microtubule organization was the same as in late pronuclear stage eggs after intact sperm injection. The first mitotic spindle was organized in eggs following ICSI with an isolated sperm head, as observed in eggs following ICSI with an intact sperm.

Conclusions: These results indicate that the MTOC is in oocyte cytoplasm during fertilization and fulfils the function when the sperm centrosome is absent. (Reprod Med Biol 2005; 4: 169–178)
Matrix remodeling and endometriosis
Authors: YANG WEI‐CHUNG VIVIAN , AU HENG‐KIEN , CHANG CHING‐WEN , CHEN HUEI‐WEN , CHEN PI‐HUA , CHEN CHIEH‐CHENG , TANG YUN‐LONG , WANG I‐TE , TZENG CHII‐RUEY
Published: May 2005

The physiological changes in endometriosis involving multiple steps of matrix remodeling include abnormal tissue growth, invasion, and adhesion formation. Endometriosis‐associated abnormal matrix remodeling is affected by several molecular factors including proteolytic enzymes and their inhibitors, which mediate tissue turnover throughout the reproductive tract to maintain the integrity of the endometrium, and ovarian steroids, which normally regulate reconstruction and breakdown of endometrium in the menstrual cycle. In addition, various growth factors, such as platelet‐derived growth factor, transform growth factor β, and epidermal growth factor, direct modulation of growth, activation, and chemotaxis which may facilitate endometrial cell adhesion onto the peritoneal mesothelium during the development of endometriosis. Furthermore, cell adhesion molecules are believed to be critically involved in most cellular‐level processes including cellular differentiation, motility, and attachment with the extracellular matrix. The present review focuses on the abnormal matrix remodeling process and its possible regulatory mechanism in association with endometriosis development. As a greater understanding of the cause of endometriosis is achieved, better treatment of the disease and its prevention become possible. (Reprod Med Biol 2005; 4: 93–99)
Activation of motility and chemotaxis in the spermatozoa: From invertebrates to humans
Authors: MORISAWA MASAAKI , YOSHIDA MANABU
Published: May 2005

Activation of the sperm motility and chemotactic behavior of sperm toward eggs are the first communication between spermatozoa and eggs at fertilization, and understanding of the phenomena is a prerequisite for progress of not only basic biology, but also clinical aspects. The nature of molecules derived from eggs by which sperm are activated and attracted towards the eggs and the molecular mechanisms underlying the sperm activation and chemotaxis have been investigated in only a few invertebrate species, sea urchins, ascidians and herring fish. However, knowledge on this phenomena has been ignored in mammalian species including humans. The current review first introduces the studies on the activation and chemotaxis of sperm in marine invertebrates, and the same phenomena in mammals including humans, are described. (Reprod Med Biol 2005; 4: 101–115)
Estrogen‐related receptor in reproductive organs
Authors: FUJIMOTO JIRO , NAKAGAWA YUMIKO , TOYOKI HIROSHI , SAKAGUCHI HIDEKI , SATO ERIKO , TAMAYA TERUHIKO
Published: May 2005

Estrogen‐related receptor (ERR) was studied in the placenta and uterine endometrium, especially endometrial cancers, among reproductive organs. In the placenta, the estrogen receptor (ER) alpha and beta mRNA levels increased from the first to the second trimester, and then decreased until normal term delivery. Estrogen‐related receptor alpha, beta and gamma mRNA levels gradually increased up to the second trimester, and then comparatively rapidly increased until normal term delivery. In endometrial cancers, ER alpha and beta mRNA levels decreased with clinical stage, myometrial invasion and dedifferentiation. Estrogen‐related receptor alpha levels increased with clinical stage and myometrial invasion, and the ERR gamma levels increased with myometrial invasion.

 Estrogen‐related receptors can bind to the steroid receptor coactivator family without any ligands, and drive transcription activity of the target genes. The manner of ERR and ER gene expressions might show a competitive interaction associated with the use of common cofactors. It is speculated that the upregulation of ERR is related to the placental growth after the downregulation of ER from the second trimester until delivery, and that ERR alpha and gamma are candidates for prognostic factors in endometrial cancer, although ERR are not directly related to tumor growth and advancement of endometrial cancer. (Reprod Med Biol 2005; 4: 129–131)
Adult motor neurons show increased susceptibility to axotomy‐induced death in mice lacking clusterin
Authors: Wicher Grzegorz K. , Aldskogius Håkan
Published: May 2005

Clusterin is a highly conserved, amphiphatic glycoprotein present in most tissues. It has been shown to be involved in the regulation of lipid transportation, clearance of cellular debris from the extracellular space and intracellular signal transduction. Clusterin is markedly up‐regulated after neural injury but the functional significance of this response is unclear. Here, we show that clusterin up‐regulation is substantially greater in hypoglossal motor neurons after hypoglossal nerve avulsion compared with nerve transection. Quantitative analyses of motor neuron numbers after the same lesions in clusterin(–/–) and clusterin(+/+) mice showed significantly larger numbers of surviving motor neurons in clusterin(+/+) mice. These results suggest that clusterin has a neuroprotective role after axotomy.
Spontaneous electrical activity and dendritic spine size in mature cerebellar Purkinje cells
Authors: Harvey Robin J. , Morando Laura , Rasetti Roberta , Strata Piergiorgio
Published: May 2005

Previous experiments have shown that in the mature cerebellum both blocking of spontaneous electrical activity and destruction of the climbing fibres by a lesion of the inferior olive have a similar profound effect on the spine distribution on the proximal dendrites of the Purkinje cells. Many new spines develop that are largely innervated by parallel fibers. Here we show that blocking electrical activity leads to a significant decrease in size of the spines on the branchlets. We have also compared the size of the spines of the proximal dendritic domain that appear during activity block and after an inferior olive lesion. In this region also, the spines in the absence of activity are significantly smaller. In the proximal dendritic domain, the new spines that develop in the absence of activity are innervated by parallel fibers and are not significantly different in size from those of the branchlets, although they are shorter. Thus, the spontaneous activity of the cerebellar cortex is necessary not only to maintain the physiological spine distribution profile in the Purkinje cell dendritic tree, but also acts as a signal that prevents spines from shrinking.
Arousal and differential Fos expression in histaminergic neurons of the ascending arousal system during a feeding‐related motivated behaviour
Authors: Valdés J. L. , Farías P. , Ocampo‐Garcés A. , Cortés N. , Serón‐Ferré M. , Torrealba F.
Published: May 2005

Arousal depends on the concerted activity of the ascending arousal system (AAS) but specific stimuli may primarily activate some nuclei of this system. Motivated behaviours are characterized by behavioural arousal, although it is not known which AAS nuclei are active during a motivated behaviour. To address this issue, rats were rendered motivated for food by fasting them for 1 day and then were enticed with food that they could not obtain for varying periods of time. We studied the level of arousal by polysomnography or radiotelemetry, and Fos‐ir in the AAS, during food enticing. We found a strong arousal and an early increase in Fos‐ir in the histaminergic neurons from the tuberomammillary nucleus, after 30 min of enticing, followed by increased Fos‐ir in the whole AAS if food enticing was prolonged to 1 or 2 hours. In contrast, food presentation to non‐motivated rats did not increase arousal or Fos‐ir in the tuberomammillary nucleus. As opposed to the active arousal of the motivated rats, passive arousal induced by sensory stimulation was associated with increased Fos‐ir in the locus coeruleus and the orexin neurons, but not with increased Fos‐ir in the tuberomammillary nucleus or in the other nuclei of the AAS. We conclude that the arousal during feeding‐related motivated behaviour is associated primarily with the activation of the tuberomammillary nucleus, while the other arousal‐related nuclei become active later on.
Selective cholinergic immunolesioning affects synaptic plasticity in developing visual cortex
Authors: Kuczewski N. , Aztiria E. , Leanza G. , Domenici L.
Published: May 2005

Cholinergic neurotransmission is known to affect activity‐dependent plasticity in various areas, including the visual cortex. However, relatively little is known about the exact role of subcortical cholinergic inputs in the regulation of plastic events in this region during early postnatal development. In the present study, synaptic transmission and plasticity in the developing visual cortex were studied following selective immunotoxic removal of the basal forebrain cholinergic afferents in 4‐day‐old rat pups. The lesion produced dramatic cholinergic neuronal and terminal fibre loss associated with decreased mRNA levels for the M1 and M2 muscarinic receptors, as well as clear‐cut impairments of long‐term potentiation (LTP) in visual cortex slices. Indeed, after theta burst stimulation of layer IV a long‐term depression (LTD) instead of an LTP was induced in immunolesioned slices. This functional change appears to be due to the lack of cholinergic input as exogenous application of acetylcholine prevented the shift from LTP to LTD. In addition, lesioned rats showed an increased sensitivity to acetylcholine (ACh). While application of 20 µm ACh produced a depression of the field potential in immunolesioned rat slices, in order to observe the same effect in control slices we had to increase ACh concentration to up to 200 µm. Taken together, our results indicate that deprivation of cholinergic input affects synaptic transmission and plasticity in developing visual cortex, suggesting that the cholinergic system could play an active role in the refinement of the cortical circuitry during maturation.
Forebrain projections to the hypothalamus are topographically organized in anurans: conservative traits as compared with amniotes
Authors: Moreno Nerea , González Agustín
Published: May 2005

The organization of the forebrain in amphibians (anamniotes) is currently being re‐evaluated in terms of evolution and several evidences have corroborated numerous traits shared by amphibians and amniotes, such as the organization of the basal ganglia and the amygdaloid complex. In the present study we have analysed the organization of forebrain afferent systems to the hypothalamus of the frog Rana perezi. In vivo and in vitro tract‐tracing techniques with dextran amines and immunohistochemistry for localizing nitric oxide synthase (NOS) in a series of single or combined experiments were used as NOS labelling reveals hypothalamic afferents arising from the lateral amygdala and the combination allowed analysis of the relationship between fibers of different origins in the same section. The results showed a large segregation of afferents in the hypothalamic region depending on their site of origin in the forebrain. Four highly topographically organized prosencephalic tracts reaching the anuran hypothalamus were observed: (i) the medial forebrain bundle, from the medial pallium and septal complex; (ii) the caudal branch of the stria terminalis formed by fibers arising in the lateral and medial amygdala; (iii) part of the lateral forebrain bundle with fibers from the central amygdala and (iv) the dorsal thalamo‐hypothalamic tract. Fibers coursing in each tract reach the hypothalamus and terminate in distinct fields. The resemblance in pattern of forebrain‐hypothalamic organization between amphibians and amniotes suggests that this feature represents an important trait conserved in the evolution of all tetrapods and therefore essential for the hypothalamic function.
Connexin26 is responsible for anionic molecule permeability in the cochlea for intercellular signalling and metabolic communications
Authors: Zhao Hong‐Bo
Published: May 2005

A gap junction is composed of two hemichannels and possesses a relatively large pore size (∼ 10–15 Å), allowing passage of ions and molecules up to 1 kDa. Here, we report that connexin hemichannels and gap junctions in the guinea pig cochlea had significant charge selectivity among permeating molecules. In coincubation with anionic and cationic fluorescent dyes, hemichannel permeability in isolated cochlear supporting cells showed significant charge selectivity; 31% of cells had only cationic dye influx and 6% of cells had only anionic dye influx. Charge‐selective influx contrary to dye size was also found, indicating charge as a dominant determinant in permeability. The cell–cell gap junctional permeability was consistent with hemichannel permeability and also showed strong charge selectivity; the permeation of anionic dyes was slower than that of cationic probes in the cochlear sensory epithelium. With a combination of immunofluorescent staining for connexin26 (Cx26) and Cx30, which are the predominant connexin isoforms in the cochlea, Cx26 was demonstrated to correlate with anionic permeability. The data indicated that cochlear gap junctions have strong charge selectivity in molecular permeability and metabolic communication. Cx26 mutation may induce specific, irreparable impairment in intercellular signalling and energy and nutrient supplies in the cochlea, causing cell degeneration and hearing loss, given that many important cell‐signalling and nutrient and energy molecules (e.g. IP3, ATP, cAMP and cGMP) are anions.
New understanding on phloem physiology and possible consequences for modelling long‐distance carbon transport
Authors: Minchin P. E. H. , Lacointe A.
Published: May 2005

Most current models of assimilate carbohydrate partitioning are based on growth patterns observed under a range of experimental conditions, from which a set of empirical rules are derived to simulate partitioning. As a result, they are not good at extrapolating to other conditions; this requires a mechanistic approach, which only transport‐resistance (TR) models currently provide. We examine an approach to incorporating recent progress in phloem physiology into the TR approach, which leads to a ‘minimalist’ Münch model of a branched system with competing sinks. In vivo whole‐plant measurements have demonstrated that C‐flow rates are dependent not only on the properties of the sink, but also on the properties of the whole transport system, and the detailed dynamics of this behaviour is mimicked by the proposed model. This model provides a sound theoretical framework for an unambiguous definition of sink and source strengths, with sink priority being an emergent property of the model. Further developments are proposed, some of which have already had limited application, to cope with the complexity of plants; the emphasis is on a modular approach, together with the importance of choosing the appropriate scale level for both structure and function. Whole‐plant experiments with in vivo measurement of the phloem dynamics will be needed to help with this choice.
Forest fragmentation effects on patch occupancy and population viability of herbaceous plant species
Authors: Honnay Olivier , Jacquemyn Hans , Bossuyt Beatrijs , Hermy Martin
Published: May 2005

Habitat fragmentation is one of the major threats to species diversity. In this review, we discuss how the genetic and demographic structure of fragmented populations of herbaceous forest plant species is affected by increased genetic drift and inbreeding, reduced mate availability, altered interactions with pollinators, and changed environmental conditions through edge effects. Reported changes in population genetic and demographic structure of fragmented plant populations have, however, not resulted in large‐scale extinction of forest plants. The main reason for this is very likely the long‐term persistence of small and isolated forest plant populations due to prolonged clonal growth and long generation times. Consequently, the persistence of small forest plant populations in a changing landscape may have resulted in an extinction debt, that is, in a distribution of forest plant species reflecting the historical landscape configuration rather than the present one. In some cases, fragmentation appears to affect ecosystem integrity rather than short‐term population viability due to the opposition of different fragmentation‐induced ecological effects. We finally discuss extinction and colonization dynamics of forest plant species at the regional scale and suggest that the use of the metapopulation concept, both because of its heuristic power and conservation applications, may be fruitful.ContentsSummary723IIntroduction724IIForest plant diversity in a historical perspective724IIIAn ecological profile of forest plant species725IVHabitat fragmentation and population viability726VEvidence of community composition changes due to habitat fragmentation?730VIAdding a regional dimension: forest plant species and metapopulation dynamics731VIIConclusions732Acknowledgements733References734