Role of P-glycoprotein in the uptake/efflux transport of oral vitamin K antagonists and rivaroxaban through the Caco-2 cell model.

Vitamin K antagonists (VKAs) are prescribed worldwide and remain the oral anticoagulant of choice. These drugs are characterized by a narrow therapeutic index and a large inter- and intra-individual variability. P-glycoprotein could contribute to this variability. The aim of this study was to investigate the involvement of P-gp in the transport of acenocoumarol, phenprocoumon and warfarin using an in vitro Caco-2 cell monolayer model. These results were compared with those obtained with rivaroxaban, a new oral anticoagulant known to be a P-gp substrate. The transport of these four drugs was assessed at pH conditions 6.8/7.4 in the presence or absence of the P-gp inhibitor cyclosporine A (10 μM) and the more potent and specific P-gp inhibitor valspodar (5 μM). Analytical quantification was performed by LC/MS. With an efflux ratio of 1.7 and a significant decrease in the efflux (Papp B-A), in the presence of P-gp inhibitors at a concentration of 50 μM, acenocoumarol can be considered as a weak P-gp substrate. Concerning phenprocoumon, the results suggest that this molecule is a poor P-gp substrate. The P-gp inhibitors did not affect significantly the transport of warfarin. The efflux of rivaroxaban was strongly inhibited by the two P-gp inhibitors. In conclusion, none of the three VKAs tested are strong P-gp substrates. However, acenocoumarol can be considered as a weak P-gp substrate and phenprocoumon as a poor P-gp substrate.

Development of fluid conduits in the auriferous shear zones of the Hutti Gold Mine, India: evidence for spatially and temporally heterogeneous fluid flow

The gold mineralization of the Hutti Mine is hosted by nine parallel, N - S trending, steeply dipping, 2 - 10 m wide shear zones, that transect Archaean amphibolites. The shear zones were formed after peak metamorphism during retrograde ductile D, shearing in the lower amphibolite facies. They were reactivated in the lower to mid greenschist facies by brittle-ductile D-3 shearing and intense quartz veining. The development of a S-2-S-3 crenulation cleavage facilitates the discrimination between the two deformation events and contemporaneous alteration and gold mineralization. Ductile D, shearing is associated with a pervasively developed distal chlorite - sed cite alteration assemblage in the outer parts of the shear zones and the proximal biotite-plagioclase alteration in the center of the shear zones. D3 is characterized by development of the inner chlorite-K-feldspar alteration, which forms a centimeter-scale alteration halo surrounding the laminated quartz veins and replaces earlier biotite along S-3. The average size of the laminated vein systems is 30-50 m along strike as well as down-dip and 2-6 m in width. Mass balance calculations suggest strong metasomatic changes for the proximal biotite-plagioclase alteration yielding mass and volume increase of ca. 16% and 12%, respectively. The calculated mass and volume changes of the distal chlorite-sericite alteration (ca. 11%, ca. 8%) are lower. The decrease in 6180 values of the whole rock from around 7.5 parts per thousand for the host rocks to 6-7 parts per thousand for the distal chlorite-sericite and the proximal biotite-plagioclase alteration and around 5 parts per thousand for the inner chlorite-K-feldspar alteration suggests hydrothermal alteration during two-stage deformation and fluid flow. The ductile D-2 deformation in the lower amphibolite facies has provided grain scale porosities by microfracturing. The pervasive, steady-state fluid flow resulted in a disseminated style of gold-sulfide mineralization and a penetrative alteration of the host rocks. Alternating ductile and brittle D3 deformation during lower to mid greenschist facies conditions followed the fault-valve process. Ductile creep in the shear zones resulted in a low permeability environment leading to fluid pressure build-up. Strongly episodic fluid advection and mass transfer was controlled by repeated seismic fracturing during the formation of laminated quartz(-gold) veins. The limitation of quartz veins to the extent of earlier shear zones indicate the importance of preexisting anisotropies for fault-valve action and economic gold mineralization. (C) 2003 Elsevier B.V. All rights reserved.

Hormonal regulation of (Na+,K+)-ATPase biosynthesis in the toad bladder. Effect of aldosterone and 3,5,3'-triiodo-L-thyronine.

Aldosterone stimulates transepithelial Na+ transport in the toad bladder, and thyroid hormone antagonizes this mineralocorticoid action. In the present study, we assessed the influence of these two hormones on the biosynthesis of (Na+,K+)ATPase, the major driving force of Na+ transport. Rates of enzyme synthesis were estimated by immunoprecipitation with monospecific alpha (96,000 daltons) and beta (60,000 daltons) subunit antibodies. After a 30-min pulse of intact tissue with [35S]methionine, the anti-alpha-serum recognized the 96,000-dalton alpha subunit and the anti-beta-serum, a 42,000-dalton protein, in total cell extracts. The biosynthesis rates of both these proteins were increased 2.8- and 2.4-fold respectively, over controls by 80 nM aldosterone after 18 h of hormone treatment. The hormonal effect was not apparent up to 3 h of incubation and was dose dependent between 0.2 and 20 nM aldosterone. The hormonal induction was antagonized by spironolactone (500-fold excess) but not by amiloride. The action of aldosterone thus seems to be a receptor-mediated process and a primary event independent of the Na+ permeability of the apical membrane. Thyroid hormone, on the other hand, had no effect on either basal or aldosterone-stimulated synthesis rates of both enzyme proteins. The results demonstrate a direct effect of aldosterone on gene expression of the (Na+,K+)-ATPase. Ultimately, this phenomenon could be linked to the late mineralocorticoid action of this hormone. On the other hand, thyroid hormone, in contrast to the situation in mammals, does not stimulate de novo enzyme synthesis in amphibia. Neither can the antimineralocorticoid action of thyroid hormone in the toad bladder be explained by an inhibition of the (Na+,K+)-ATPase synthesis.

Prevalence and risk factors for Leptospira spp. in cattle herds in the south central region of Paraná state

The aim of this study was to determine the prevalence of anti-Leptospira spp. antibodies and the risk factors for Leptospira spp. infection in breeding cattle herds in the south central region of Paraná state. It was based on the statistic delineation/serological samples and information regarding the selected farms employed in the study of bovine brucellosis for Paraná state in the context of National Program for Control and Eradication of Brucellosis and Tuberculosis. A total of 1.880 females aged >24 months from 274 non vaccinated herds were studied. Serum samples were tested for antibodies against Leptospira spp. using microscopic agglutination test (MAT) with 22 Leptospira serovars. The epidemiological questionnaire was applied on all the selected farms and aimed to obtain epidemiological data. Hundred eighty one of 274 herds were positive for Leptospira spp./presenting prevalence of positive herds of 66.06% (IC95%=60.12-71,65%). Presence of >43 cattle (OR=3.120; IC=1.418-6.867)/animal purchase (OR=2.010; IC=1.154-3.500)/rent of pastures (OR=2.925; IC=1.060-8.068) and presence of maternity paddock (OR=1.981; IC=1,068-3,676) were identified as risk factors for leptospirosis due to any serovar in the multivariate logistic regression. Risk factors for leptospirosis due to serovar Hardjo were presence of >43 cattle (OR=3.622; IC=1.512-8,677)/animal purchase (OR=3.143; IC=1.557-6.342)/rent of pastures (OR=4.070; IC=1.370-12.087) and presence of horses (OR=2.981; IC=1.321-6.726). These results indicate that Leptospira spp. infection is widespread in the south central region of Paraná state and that factors related to the herd characteristic and management are associated with the infection.

The role of B-mode ultrasonography in the musculoskeletal anatomical evaluation of the cervical region of the dog spine

This study characterized the normal musculoskeletal anatomy of the cervical segment of the spine of dogs by means of B-mode ultrasonography. The objective was to establish the role of B-mode ultrasonography for the anatomical evaluation of the cervical spine segment in dogs, by comparing the ultrasonographic findings with images by computed tomography and magnetic resonance imaging. The ultrasound examination, in transverse and median sagittal sections, allowed to identify a part of the epaxial cervical musculature, the bone surface of the cervical vertebrae and parts of the spinal cord through restricted areas with natural acoustic windows, such as between the atlanto-occipital joint, axis and atlas, and axis and the third cervical vertebra. The images, on transverse and sagittal planes, by low-field magnetic resonance imaging, were superior for the anatomical identification of the structures, due to higher contrast between the different tissues in this modality. Computed tomography showed superiority for bone detailing when compared with ultrasonography. As for magnetic resonance imaging, in addition to the muscles and cervical vertebrae, it is possible to identify the cerebrospinal fluid and differentiate between the nucleus pulposus and annulus fibrosus of the intervertebral discs. Although not the scope of this study, with knowledge of the ultrasonographic anatomy of this region, it is believed that some lesions can be identified, yet in a limited manner, when compared with the information obtained mainly with magnetic resonance imaging. The ultrasound examination presented lower morphology diagnostic value compared with the other modalities.

Opiatergic participation in the thirst-inhibiting effect of acute third ventricle injections of cadmium (Cd2+) and lead (Pb2+)

We have previously demonstrated that acute third ventricle injections of both lead and cadmium prevent the dipsogenic response elicited by dehydration or by central injections of dipsogenic agents such as angiotensin II, carbachol and isoproterenol in rats. We have also shown that the antidipsogenic action of cadmium may be due, at least in part, to activation of thirst-inhibitory central serotonergic pathways. In the present paper we show that in Wistar male rats the antidipsogenic effect of both lead acetate (3.0 nmol/rat) and cadmium chloride (3.0 nmol/rat) may be partially dependent on the activation of brain opiatergic pathways since central injections of naloxone (82.5 nmol/rat), a non-selective opioid antagonist, blunt the thirst-inhibiting effect of these metals. One hundred and twenty minutes after the second third ventricle injections, dehydrated animals (14 h overnight) receiving saline + sodium acetate displayed a high water intake (7.90 ± 0.47 ml/100 g body weight) whereas animals receiving saline + lead acetate drank 3.24 ± 0.47 ml/100 g body weight. Animals receiving naloxone + lead acetate drank 6.94 ± 0.60 ml/100 g body weight. Animals receiving saline + saline drank 8.16 ± 0.66 ml/100 g body weight whilst animals receiving saline + cadmium chloride drank 1.63 ± 0.37 ml/100 g body weight. Animals receiving naloxone + cadmium chloride drank 8.01 ± 0.94 ml/100 g body weight. It is suggested that acute third ventricle injections of both lead and cadmium exert their antidipsogenic effect by activating thirst-inhibiting opioid pathways in the brain.

The bradycardic and hypotensive responses to serotonin are reduced by activation of GABA A receptors in the nucleus tractus solitarius of awake rats

We investigated the effects of bilateral injections of the GABA receptor agonists muscimol (GABA A) and baclofen (GABA B) into the nucleus tractus solitarius (NTS) on the bradycardia and hypotension induced by iv serotonin injections (5-HT, 2 µg/rat) in awake male Holtzman rats. 5-HT was injected in rats with stainless steel cannulas implanted bilaterally in the NTS, before and 5, 15, and 60 min after bilateral injections of muscimol or baclofen into the NTS. The responses to 5-HT were tested before and after the injection of atropine methyl bromide. Muscimol (50 pmol/50 nl, N = 8) into the NTS increased basal mean arterial pressure (MAP) from 115 ± 4 to 144 ± 6 mmHg, did not change basal heart rate (HR) and reduced the bradycardia (-40 ± 14 and -73 ± 26 bpm at 5 and 15 min, respectively, vs -180 ± 20 bpm for the control) and hypotension (-11 ± 4 and -14 ± 4 mmHg, vs -40 ± 9 mmHg for the control) elicited by 5-HT. Baclofen (12.5 pmol/50 nl, N = 7) into the NTS also increased basal MAP, but did not change basal HR, bradycardia or hypotension in response to 5-HT injections. Atropine methyl bromide (1 mg/kg body weight) injected iv reduced the bradycardic and hypotensive responses to 5-HT injections. The stimulation of GABA A receptors in the NTS of awake rats elicits a significant increase in basal MAP and decreases the cardiac Bezold-Jarisch reflex responses to iv 5-HT injections.

Análisis de una iniciativa estudiantil de investigación surgida en Criptogamia, una asignatura optativa de segundo ciclo de Biología = Analysis of a students’ research initiative originating in Cryptogamy, an elective course in the last two years of the Biology degree

Presentamos una experiencia exitosa de aprendizaje que partió de Criptogamia (asignatura optativa de segundo ciclo de Biología), que dio lugar a un proyecto de investigación gestionado por los propios alumnos. La iniciativa se consolidó estableciendo una Asociación de Estudiantes centrada en investigación y divulgación. En poco tiempo, los participantes han presentado comunicaciones científicas, y organizado actividades dirigidas a diversos públicos, dentro y fuera de la comunidad universitaria. Actualmente se plantea una colaboración multidisciplinar con otros organismos de investigación y la extensión de su ámbito de estudio. Abordamos su incidencia en el aprendizaje en varios aspectos: científico (técnicas específicas, rigor, búsqueda de información e interpretación de resultados), comunicativo (estructuración y presentación de la información obtenida, para diversos públicos), y organizativo, incluyendo el trabajo en equipo. Aunque de carácter espontáneo, esta experiencia muestra rasgos evaluables en cuanto a sus posibilidades para otras asignaturas. Analizamos las características y planteamiento de esta optativa, el perfil de sus alumnos, y el contexto universitario que la acoge. Detectamos como factores principales los aspectos participativos de la asignatura, la cohesión del grupo, el carácter voluntario de la implicación, los beneficios percibidos por los estudiantes, y la disponibilidad de recursos humanos (supervisión) y materiales (equipamiento y subvenciones)

In situ degradability of dry matter and neutral-detergent fibre of thorn scrubland forage consumed by goats in the semi-arid region of north Mexico

Three goats provided with oesophageal and ruminal cannulae were used to determine variations in dry matter (DM) and neutral-detergent fibre (NDF) degradability of the forage consumed when grazing thorn scrubland in the semi-arid region of north Mexico, during two consecutive dry and wet periods. Ingesta samples were incubated intraruminally, the data were fitted to the exponential equation P = a + b (1-e(-ct)) and statistically analysed using a randomized-block design. Organic matter and crude protein (CP) contents were higher (P < 0.05) in the wet seasons. Values of NDF were similar in dry and wet season of both years whereas higher numerical values of acid-detergent fibre (ADF), lignin and cellulose were registered in the dry seasons. DM and NDF degradabilities after 24 and 48 h of ruminal incubation were higher (P < 0.05) in the wet seasons. Higher values (P < 0.05) in DM and NDF bag losses at zero time (A fraction) were registered in the two wet seasons. The insoluble but fermentable DM and NDF (B fractions) were higher (P < 0.05) in the 1999 wet season and variable in the rest of the studied period. Numerically higher values of DM and NDF c fraction were found in wet periods, whereas DM and NDF potential degradabilities were higher (P < 0.05) in the wet season in 1999 and similar across seasons in 2000. Lowest (P < 0.05) contents of CP in grazed forage, DM and NDF degradabilities after 48 h of ruminal incubation, and A, and B, and c fractions were observed in the dry seasons. Thus, these results may be related to both the lower feeding value of forage consumed by the animals and lower performance of livestock during this period. Then, the DM and NDF degradability after 48 h, together with the insoluble but fermentable matter and the c fraction permit the nutritive value of the forage consumed by grazing goats to be accurately described.

Chemical composition and in vitro ruminal fermentation of common tree forages in the semi-arid rangelands of Swaziland

Browse plants play an important role in providing feed for livestock in semi-arid rangelands of Africa. Chemical composition and in vitro ruminal fermentation of leaves collected from Acacia burkei, Acacia tortilis, Acacia nilotica, Dichrostachys cinerea and Ehretia obtusifolia in communal grazing lands in the lowveld of Swaziland is presented. Leaves were collected from trees located on two soil types (i.e., lithosol and vertisol) in the communal land but it had no effect on the chemical composition of tree leaves. The NDFom and ADFom content were highest in D. cinerea and A. burkei and lowest in E. obtusifolia and A. nilotica. Crude protein (CP) contents ranged between 108 g/kg and 122 g/kg DM. D. cinerea had the highest Ca and Mg content, while A. tortilis had the lowest. There were marked variations in K level amongst browse species, with A. tortilis (9.1 g/kg DM) having the highest value. The P, Zn and Fe did not differ between browse species. Soil type and tree species interaction impacted in vitro fermentation parameters. Extent of fermentation, as measured by 48 h cumulative gas production, and organic matter degradability was highest in E. obtusifolia leaves and lowest in D. cinerea leaves within soil type. Fermentation efficiency, as measured by partitioning factors, was highest in A. nilotica leaves. Leaves of E. obtusifolia could be a valuable supplementary feedstuff for ruminant livestock due to its in vitro fermentation characteristics as well as low fibre and moderate CP levels. (c) 2007 Elsevier B.V. All rights reserved.

The effects of environmental perturbation and measurement error on estimates of the shape parameter in the theta-logistic model of population regulation

The theta-logistic is a widely used generalisation of the logistic model of regulated biological processes which is used in particular to model population regulation. Then the parameter theta gives the shape of the relationship between per-capita population growth rate and population size. Estimation of theta from population counts is however subject to bias, particularly when there are measurement errors. Here we identify factors disposing towards accurate estimation of theta by simulation of populations regulated according to the theta-logistic model. Factors investigated were measurement error, environmental perturbation and length of time series. Large measurement errors bias estimates of theta towards zero. Where estimated theta is close to zero, the estimated annual return rate may help resolve whether this is due to bias. Environmental perturbations help yield unbiased estimates of theta. Where environmental perturbations are large, estimates of theta are likely to be reliable even when measurement errors are also large. By contrast where the environment is relatively constant, unbiased estimates of theta can only be obtained if populations are counted precisely Our results have practical conclusions for the design of long-term population surveys. Estimation of the precision of population counts would be valuable, and could be achieved in practice by repeating counts in at least some years. Increasing the length of time series beyond ten or 20 years yields only small benefits. if populations are measured with appropriate accuracy, given the level of environmental perturbation, unbiased estimates can be obtained from relatively short censuses. These conclusions are optimistic for estimation of theta. (C) 2008 Elsevier B.V All rights reserved.

Neurokinin B is a paracrine vasodilator in the human fetal placental circulation

Neurokinin (NK) B is a member of the tachykinin family of neurotransmitters, exerting hypotensive or hypertensive effects in the mammalian vasculature through synaptic release from peripheral neurons, according to either NK1 and NK2 or NK3 receptor subtype expression, respectively. There is recent evidence that NKB is expressed by the syncytiotrophoblast of the human placenta, an organ that is not innervated. We hypothesized that NKB is a paracrine modulator of tone in the fetal placental circulation. We tested this hypothesis using the in vitro perfused human placental cotyledon. Our data show that NKB is a dilator of the fetal vasculature, causing a maximal 25.1+/-4.5% (mean+/-SEM; n=5) decrease in fetal-side arterial hydrostatic pressure (5-muM NKB bolus; effective concentration in the circulation, 1.89 nM) after preconstriction with U-46619. RT-PCR demonstrated the presence of mRNA for NK1 and NK2 tachykinin receptors in the placenta. Using selective receptor antagonists, we found that NKB-induced vasodilation is through the NK1 receptor subtype. We found no evidence for the involvement of either nitric oxide or prostacyclin in this response. This study demonstrates a paracrine role for NKB in the regulation of fetal placental vascular tone.

Glycogen synthase kinase 3 (GSK3) in the heart: a point of integration in hypertrophic signalling and a therapeutic target? A critical analysis

Glycogen synthase kinase 3 (GSK3, of which there are two isoforms, GSK3alpha and GSK3beta) was originally characterized in the context of regulation of glycogen metabolism, though it is now known to regulate many other cellular processes. Phosphorylation of GSK3alpha(Ser21) and GSK3beta(Ser9) inhibits their activity. In the heart, emphasis has been placed particularly on GSK3beta, rather than GSK3alpha. Importantly, catalytically-active GSK3 generally restrains gene expression and, in the heart, catalytically-active GSK3 has been implicated in anti-hypertrophic signalling. Inhibition of GSK3 results in changes in the activities of transcription and translation factors in the heart and promotes hypertrophic responses, and it is generally assumed that signal transduction from hypertrophic stimuli to GSK3 passes primarily through protein kinase B/Akt (PKB/Akt). However, recent data suggest that the situation is far more complex. We review evidence pertaining to the role of GSK3 in the myocardium and discuss effects of genetic manipulation of GSK3 activity in vivo. We also discuss the signalling pathways potentially regulating GSK3 activity and propose that, depending on the stimulus, phosphorylation of GSK3 is independent of PKB/Akt. Potential GSK3 substrates studied in relation to myocardial hypertrophy include nuclear factors of activated T cells, beta-catenin, GATA4, myocardin, CREB, and eukaryotic initiation factor 2Bvarepsilon. These and other transcription factor substrates putatively important in the heart are considered. We discuss whether cardiac pathologies could be treated by therapeutic intervention at the GSK3 level but conclude that any intervention would be premature without greater understanding of the precise role of GSK3 in cardiac processes.

Pre-Columbian landscape impact and agriculture in the Monumental Mound region of the Llanos de Moxos, lowland Bolivia

We present a multiproxy study of land use by a pre-Columbian earth mounds culture in the Bolivian Amazon. The Monumental Mounds Region (MMR) is an archaeological sub-region characterized by hundreds of pre-Columbian habitation mounds associated with a complex network of canals and causeways, and situated in the forest–savanna mosaic of the Llanos de Moxos. Pollen, phytolith, and charcoal analyses were performed on a sediment core from a large lake (14 km2), Laguna San José (14°56.97′S, 64°29.70′W).We found evidence of high levels of anthropogenic burning from AD 400 to AD 1280, corroborating dated occupation layers in two nearby excavated habitation mounds. The charcoal decline pre-dates the arrival of Europeans by at least 100 yr, and challenges the notion that the mounds culture declined because of European colonization. We show that the surrounding savanna soils were sufficiently fertile to support crops, and the presence of maize throughout the record shows that the area was continuously cultivated despite land-use change at the end of the earthmounds culture. We suggest that burning was largely confined to the savannas, rather than forests, and that pre-Columbian deforestation was localized to the vicinity of individual habitation mounds, whereas the inter-mound areas remained largely forested.