Metallothionein I and II gene expression as a response to metal contamination in bank vole Clethrionomys glareolus

The expression of two metallothionein genes (Mt-I and Mt-II) in the liver, kidney, and gonad of bank voles collected at four metal-contaminated sites (Cd, Zn, Pb, and Fe) were measured using the quantitative real-time PCR method (QPCR). Relative Mt gene expression was calculated by applying a normalization factor (NF) using the expression of two housekeeping genes, ribosomal 18S and beta-actin. Relative Mt expression in tissues of animals from contaminated sites was up to 54.8-fold higher than those from the reference site for Mt-I and up to 91.6-fold higher for Mt-II. Mt-II gene expression in the livers of bank voles from contaminated sites was higher than Mt-I gene expression. Inversely, Mt-II expression in the kidneys of voles was lower than Mt-I expression. Positive correlations between cadmium levels in the tissues and Mt-I were obtained in all studied tissues. Zinc, which undergoes homeostatic regulation, correlated positively with both Mt-I and Mt-II gene expression only in the kidney. Results showed that animals living in chronically contaminated environments intensively activate detoxifying mechanisms such as metallothionein expression. This is the first time that QPCR techniques to measure MT gene expression have been applied to assess the impact of environmental metal pollution on field collected bank voles.

Mechanisms of inverse agonist action at D-2 dopamine receptors

1 Mechanisms of inverse agonist action at the D-2(short) dopamine receptor have been examined. 2 Discrimination of G-protein-coupled and -uncoupled forms of the receptor by inverse agonists was examined in competition ligand-binding studies versus the agonist [H-3]NPA at a concentration labelling both G-protein-coupled and -uncoupled receptors. 3 Competition of inverse agonists versus [H-3] NPA gave data that were fitted best by a two-binding site model in the absence of GTP but by a one-binding site model in the presence of GTP. K-i values were derived from the competition data for binding of the inverse agonists to G-protein-uncoupled and -coupled receptors. K-coupled and K-uncoupled were statistically different for the set of compounds tested ( ANOVA) but the individual values were different in a post hoc test only for (+)-butaclamol. 4 These observations were supported by simulations of these competition experiments according to the extended ternary complex model. 5 Inverse agonist efficacy of the ligands was assessed from their ability to reduce agonist-independent [S-35]GTPγ S binding to varying degrees in concentration-response curves. Inverse agonism by (+)-butaclamol and spiperone occurred at higher potency when GDP was added to assays, whereas the potency of (-)-sulpiride was unaffected. 6 These data show that some inverse agonists ((+)-butaclamol, spiperone) achieve inverse agonism by stabilising the uncoupled form of the receptor at the expense of the coupled form. For other compounds tested, we were unable to define the mechanism.

Mechanisms and function of flower and inflorescence reversion

Flower and inflorescence reversion involve a switch from floral development back to vegetative development, thus rendering flowering a phase in an ongoing growth pattern rather than a terminal act of the meristem. Although it can be considered an unusual event, reversion raises questions about the nature and function of flowering. It is linked to environmental conditions and is most often a response to conditions opposite to those that induce flowering. Research on molecular genetic mechanisms underlying plant development over the last 15 years has pinpointed some of the key genes involved in the transition to flowering and flower development. Such investigations have also uncovered mutations which reduce floral maintenance or alter the balance between vegetative and floral features of the plant. How this information contributes to an understanding of floral reversion is assessed here. One issue that arises is whether floral commitment (defined as the ability to continue flowering when inductive conditions no longer exist) is a developmental switch affecting the whole plant or is a mechanism which assigns autonomy to individual meristems. A related question is whether floral or vegetative development is the underlying default pathway of the plant. This review begins by considering how studies of flowering in Arabidopsis thaliana have aided understanding of mechanisms of floral maintenance. Arabidopsis has not been found to revert to leaf production in any of the conditions or genetic backgrounds analysed to date. A clear-cut reversion to leaf production has, however, been described in Impatiens balsamina. It is proposed that a single gene controls whether Impatiens reverts or can maintain flowering when inductive conditions are removed, and it is inferred that this gene functions to control the synthesis or transport of a leaf-generated signal. But it is also argued that the susceptibility of Impatiens to reversion is a consequence of the meristem-based mechanisms controlling development of the flower in this species. Thus, in Impatiens, a leaf-derived signal is critical for completion of flowering and can be considered to be the basis of a plant-wide floral commitment that is achieved without accompanying meristem autonomy. The evidence, derived from in vitro and other studies, that similar mechanisms operate in other species is assessed. It is concluded that most species (including Arabidopsis) are less prone to reversion because signals from the leaf are less ephemeral, and the pathways driving flower development have a high level of redundancy that generates meristem autonomy even when leaf-derived signals are weak. This gives stability to the flowering process, even where its initiation is dependent on environmental cues. On this interpretation, Impatiens reversion appears as an anomaly resulting from an unusual combination of leaf signalling and meristem regulation. Nevertheless, it is shown that the ability to revert can serve a function in the life history strategy (perenniality) or reproductive habit (pseudovivipary) of many plants. In these instances reversion has been assimilated into regular plant development and plays a crucial role there.

Mechanisms of agonist action at D-2 dopamine receptors

In this study, we investigated the biochemical mechanisms of agonist action at the G protein-coupled D-2 dopamine receptor expressed in Chinese hamster ovary cells. Stimulation of guanosine 5'-O-(3-[S-35]thio) triphosphate ([S-35]GTPgammaS) binding by full and partial agonists was determined at different concentrations of [S-35]GTPgammaS (0.1 and 10 nM) and in the presence of different concentrations of GDP. At both concentrations of [S-35]GTPgammaS, increasing GDP decreased the [S-35]GTPgammaS binding observed with maximally stimulating concentrations of agonist, with partial agonists exhibiting greater sensitivity to the effects of GDP than full agonists. The relative efficacy of partial agonists was greater at the lower GDP concentrations. Concentration-response experiments were performed for a range of agonists at the two [S-35]GTPgammaS concentrations and with different concentrations of GDP. At 0.1 nM [S-35]GTPgammaS, the potency of both full and partial agonists was dependent on the GDP concentration in the assays. At 10 nM [S-35]GTPgammaS, the potency of full agonists exhibited a greater dependence on the GDP concentration, whereas the potency of partial agonists was virtually independent of GDP. We concluded that at the lower [S-35]GTPgammaS concentration, the rate-determining step in G protein activation is the binding of [S-35]GTPgammaS to the G protein. At the higher [S-35]GTPgammaS concentration, for full agonists, [S-35]GTPgammaS binding remains the slowest step, whereas for partial agonists, another (GDP-independent) step, probably ternary complex breakdown, becomes rate-determining.

Rate-limiting steps in the development of atherosclerosis: the response-to-influx theory

A large number of processes are involved in the pathogenesis of atherosclerosis but it is unclear which of them play a rate-limiting role. One way of resolving this problem is to investigate the highly non-uniform distribution of disease within the arterial system; critical steps in lesion development should be revealed by identifying arterial properties that differ between susceptible and protected sites. Although the localisation of atherosclerotic lesions has been investigated intensively over much of the 20th century, this review argues that the factor determining the distribution of human disease has only recently been identified. Recognition that the distribution changes with age has, for the first time, allowed it to be explained by variation in transport properties of the arterial wall; hitherto, this view could only be applied to experimental atherosclerosis in animals. The newly discovered transport variations which appear to play a critical role in the development of adult disease have underlying mechanisms that differ from those elucidated for the transport variations relevant to experimental atherosclerosis: they depend on endogenous NO synthesis and on blood flow. Manipulation of transport properties might have therapeutic potential. Copyright (C) 2004 S. Karger AG, Basel.

Identification of hepatic molecular mechanisms of action of alpha-tocopherol using global gene expression profile analysis in rats

The recent discovery that vitamin E (VE) regulates gene activity at the transcriptional level indicates that VE may exert part of its biological effects by mechanisms which may be independent of its well-recognised antioxidant function. The objective of this study was the identification of hepatic vitamin E-sensitive genes and examination of the effects of VE on their corresponding biological endpoints. Two groups of male rats were randomly assigned to either a VE-sufficient diet or to a control diet deficient in VE for 290 days. High-density oligonucleotide microarrays comprising over 7000 genes were used to assess the transcriptional response of the liver. Differential gene expression was monitored over a period of 9 months, at four different time-points, and rats were individually profiled. This experimental strategy identified several VE-sensitive genes, which were chronically altered by dietary VE. VE supplementation down-regulated scavenger receptor CD36, coagulation factor IX and 5-alpha-steroid reductase type 1 mRNA levels while hepatic gamma glutamyl-cysteinyl synthetase was significantly up-regulated. Measurement of the corresponding biological endpoints such as activated partial thromboplastin time, plasma dihydrotestosterone and hepatic glutathione substantiated the gene chip data which indicated that dietary VE plays an important role in a range of metabolic processes within the liver. (C) 2004 Elsevier B.V. All rights reserved.

Mechanisms underlying agonist efficacy

Agonist efficacy is a measure of how well an agonist can stimulate a response system linked to a receptor. Efficacy can be assessed in functional assays and various parameters (E-max, K-A/EC50, E-max center dot K-A/EC50) determined. The E-max center dot K-A/EC50 parameter provides a good estimate of efficacy across the full range of efficacy. A convenient assay for the efficacy of agonists for some receptors is provided by the [S-35]GTP[S] (guanosine 5'-[gamma-[S-35]thio]triphosphate)-binding assay. in this assay, the normal GTP-binding event in GPCR (G-protein-coupled receptor) activation is replaced by the binding of the non-hydrolysable analogue [S-35]GTP[S]. This assay may be used to profile ligands for their efficacy, and an example here is the D-2 dopamine receptor where an efficacy scale has been set up using this assay. The mechanisms underlying the assay have been probed. The time course of [S-35]GTP[S] binding follows a pseudo-first-order reaction with [S-35]GTP[S] binding reaching equilibrium after approx. 3 h. The [S-35]GTP[S]-binding event is the rate-deter mining step in the assay. Agonists regulate the maximal level of [S-35]GTP[S] bound, rather than the rate constant for binding. The [S-35]GTP[S]-binding assay therefore determines agonist efficacy on the basis of the amount of [S-35]GTP[S] bound rather than the rate of binding.

Mechanisms of inverse agonist action at D-2 dopamine receptors

1 Mechanisms of inverse agonist action at the D-2(short) dopamine receptor have been examined. 2 Discrimination of G-protein-coupled and -uncoupled forms of the receptor by inverse agonists was examined in competition ligand-binding studies versus the agonist [H-3]NPA at a concentration labelling both G-protein-coupled and -uncoupled receptors. 3 Competition of inverse agonists versus [H-3] NPA gave data that were fitted best by a two-binding site model in the absence of GTP but by a one-binding site model in the presence of GTP. K-i values were derived from the competition data for binding of the inverse agonists to G-protein-uncoupled and -coupled receptors. K-coupled and K-uncoupled were statistically different for the set of compounds tested ( ANOVA) but the individual values were different in a post hoc test only for (+)-butaclamol. 4 These observations were supported by simulations of these competition experiments according to the extended ternary complex model. 5 Inverse agonist efficacy of the ligands was assessed from their ability to reduce agonist-independent [S-35]GTPγ S binding to varying degrees in concentration-response curves. Inverse agonism by (+)-butaclamol and spiperone occurred at higher potency when GDP was added to assays, whereas the potency of (-)-sulpiride was unaffected. 6 These data show that some inverse agonists ((+)-butaclamol, spiperone) achieve inverse agonism by stabilising the uncoupled form of the receptor at the expense of the coupled form. For other compounds tested, we were unable to define the mechanism.

ERP correlates of shared control mechanisms involved in saccade preparation and in covert attention

We investigated whether attention shifts and eye movement preparation are mediated by shared control mechanisms, as claimed by the premotor theory of attention. ERPs were recorded in three tasks where directional cues presented at the beginning of each trial instructed participants to direct their attention to the cued side without eye movements (Covert task), to prepare an eye movement in the cued direction without attention shifts (Saccade task) or both (Combined task). A peripheral visual Go/Nogo stimulus that was presented 800 ms after cue onset signalled whether responses had to be executed or withheld. Lateralised ERP components triggered during the cue–target interval, which are assumed to reflect preparatory control mechanisms that mediate attentional orienting, were very similar across tasks. They were also present in the Saccade task, which was designed to discourage any concomitant covert attention shifts. These results support the hypothesis that saccade preparation and attentional orienting are implemented by common control structures. There were however systematic differences in the impact of eye movement programming and covert attention on ERPs triggered in response to visual stimuli at cued versus uncued locations. It is concluded that, although the preparatory processes underlying saccade programming and covert attentional orienting may be based on common mechanisms, they nevertheless differ in their spatially specific effects on visual information processing.

Iron homeostasis and management of oxidative stress response in bacteria

Iron is both an essential nutrient for the growth of microorganisms, as well as a dangerous metal due to its capacity to generate reactive oxygen species (ROS) via the Fenton reaction. For these reasons, bacteria must tightly control the uptake and storage of iron in a manner that restricts the build-up of ROS. Therefore, it is not surprising to find that the control of iron homeostasis and responses to oxidative stress are coordinated. The mechanisms concerned with these processes, and the interactions involved, are the subject of this review.

Turning on the alarm: the neural mechanisms of the transition from innocuous to painful sensation

The experience of pain occurs when the level of a stimulus is sufficient to elicit a marked affective response, putatively to warn the organism of potential danger and motivate appropriate behavioral responses. Understanding the biological mechanisms of the transition from innocuous to painful levels of sensation is essential to understanding pain perception as well as clinical conditions characterized by abnormal relationships between stimulation and pain response. Thus, the primary objective of this study was to characterize the neural response associated with this transition and the correspondence between that response and subjective reports of pain. Towards this goal, this study examined BOLD response profiles across a range of temperatures spanning the pain threshold. 14 healthy adults underwent functional magnetic resonance imaging (fMRI) while a range of thermal stimuli (44-49oC) were applied. BOLD responses showed a sigmoidal profile along the range of temperatures in a network of brain regions including insula and mid- cingulate, as well as a number of regions associated with motor responses including ventral lateral nuclei of the thalamus, globus pallidus and premotor cortex. A sigmoid function fit to the BOLD responses in these regions explained up to 85% of the variance in individual pain ratings, and yielded an estimate of the temperature of steepest transition from non-painful to painful heat that was nearly identical to that generated by subjective ratings. These results demonstrate a precise characterization of the relationship between objective levels of stimulation, resulting neural activation, and subjective experience of pain and provide direct evidence for a neural mechanism supporting the nonlinear transition from innocuous to painful levels along the sensory continuum.

The role of atmosphere feedbacks during ENSO in the CMIP3 models. Part II: using AMIP runs to understand the heat flux feedback mechanisms

Several studies using ocean–atmosphere general circulation models (GCMs) suggest that the atmospheric component plays a dominant role in the modelled El Niño-Southern Oscillation (ENSO). To help elucidate these findings, the two main atmosphere feedbacks relevant to ENSO, the Bjerknes positive feedback (μ) and the heat flux negative feedback (α), are here analysed in nine AMIP runs of the CMIP3 multimodel dataset. We find that these models generally have improved feedbacks compared to the coupled runs which were analysed in part I of this study. The Bjerknes feedback, μ, is increased in most AMIP runs compared to the coupled run counterparts, and exhibits both positive and negative biases with respect to ERA40. As in the coupled runs, the shortwave and latent heat flux feedbacks are the two dominant components of α in the AMIP runs. We investigate the mechanisms behind these two important feedbacks, in particular focusing on the strong 1997–1998 El Niño. Biases in the shortwave flux feedback, α SW, are the main source of model uncertainty in α. Most models do not successfully represent the negative αSW in the East Pacific, primarily due to an overly strong low-cloud positive feedback in the far eastern Pacific. Biases in the cloud response to dynamical changes dominate the modelled α SW biases, though errors in the large-scale circulation response to sea surface temperature (SST) forcing also play a role. Analysis of the cloud radiative forcing in the East Pacific reveals model biases in low cloud amount and optical thickness which may affect α SW. We further show that the negative latent heat flux feedback, α LH, exhibits less diversity than α SW and is primarily driven by variations in the near-surface specific humidity difference. However, biases in both the near-surface wind speed and humidity response to SST forcing can explain the inter-model αLH differences.

Mechanisms linking volcanic aerosols to the Atlantic meridional overturning circulation

This study examines the sensitivity of the climate system to volcanic aerosol forcing in the third climate configuration of the Met Office Unified Model (HadCM3). The main test case was based on the 1880s when there were several volcanic eruptions, the well-known Krakatau being the largest. These eruptions increased atmospheric aerosol concentrations and induced a period of global cooling surface temperatures. In this study, an ensemble of HadCM3 has been integrated with the standard set of radiative forcings and aerosols from the Intergovernmental Panel on Climate Change Fourth Assessment Report simulations, from 1860 to present. A second ensemble removes the volcanic aerosols from 1880 to 1899. The all-forcings ensemble shows an attributable 1.2-Sv (1 Sv ≡ 106 m3 s−1) increase in the Atlantic meridional overturning circulation (AMOC) at 45°N—with a 0.04-PW increase in meridional heat transport at 40°N and increased northern Atlantic SSTs—starting around 1894, approximately 11 years after the first eruption, and lasting a further 10 years at least. The mechanisms responsible are traced to the Arctic, with suppression of the global water cycle (high-latitude precipitation), which leads to an increase in upper-level Arctic and Greenland Sea salinities. This then leads to increased convection in the Greenland–Iceland–Norwegian (GIN) Seas, enhanced Denmark Strait overflows, and AMOC changes with density anomalies traceable southward along the western Atlantic boundary. The authors investigate whether a similar response to the Pinatubo eruption in 1991 could still be ongoing, but do not find strong evidence.

Dynamics of the lower stratospheric circulation response to ENSO

A robust feature of the observed response to El Nin˜o–Southern Oscillation (ENSO) is an altered circulation in the lower stratosphere. When sea surface temperatures (SSTs) in the tropical Pacific are warmer there is enhanced upwelling and cooling in the tropical lower stratosphere and downwelling and warming in the midlatitudes, while the opposite is true of cooler SSTs. The midlatitude lower stratospheric response to ENSO is larger in the Southern Hemisphere (SH) than in the Northern Hemisphere (NH). In this study the dynamical version of the Canadian Middle Atmosphere Model (CMAM) is used to simulate 25 realizations of the atmospheric response to the 1982/83 El Nin˜o and the 1973/74 La Nin˜ a. This version ofCMAMis a comprehensive high-top general circulation model that does not include interactive chemistry. The observed lower stratospheric response to ENSO is well reproduced by the simulations, allowing them to be used to investigate the mechanisms involved. Both the observed and simulated responses maximize in December–March and so this study focuses on understanding the mechanisms involved in that season. The response in tropical upwelling is predominantly driven by anomalous transient synoptic-scale wave drag in the SH subtropical lower stratosphere, which is also responsible for the compensating SH midlatitude response. This altered wave drag stems from an altered upward flux of wave activity from the troposphere into the lower stratosphere between 208 and 408S. The altered flux of wave activity can be divided into two distinct components. In the Pacific, the acceleration of the zonal wind in the subtropics from the warmer tropical SSTs results in a region between the midlatitude and subtropical jets where there is an enhanced source of low phase speed eddies. At other longitudes, an equatorward shift of the midlatitude jet from the extratropical tropospheric response to El Nin˜o results in an enhanced source of waves of higher phase speeds in the subtropics. The altered resolved wave drag is only apparent in the SH and the difference between the two hemispheres can be related to the difference in the climatological jet structures in this season and the projection of the wind anomalies associated with ENSO onto those structures.

Response of the Atlantic meridional overturning circulation to a reversal of greenhouse gas increases

The reversibility of the Atlantic meridional overturning circulation (AMOC) is investigated in multi-model experiments using global climate models (GCMs) where CO2 concentrations are increased by 1 or 2 % per annum to 2× or 4× preindustrial conditions. After a period of stabilisation the CO2 is decreased back to preindustrial conditions. In most experiments when the CO2 decreases, the AMOC recovers before becoming anomalously strong. This "overshoot" is up to an extra 18.2Sv or 104 % of its preindustrial strength, and the period with an anomalously strong AMOC can last for several hundred years. The magnitude of this overshoot is shown to be related to the build up of salinity in the subtropical Atlantic during the previous period of high CO2 levels. The magnitude of this build up is partly related to anthropogenic changes in the hydrological cycle. The mechanisms linking the subtropical salinity increase to the subsequent overshoot are analysed, supporting the relationship found. This understanding is used to explain differences seen in some models and scenarios. In one experiment there is no overshoot because there is little salinity build up, partly as a result of model differences in the hydrological cycle response to increased CO2 levels and partly because of a less aggressive scenario. Another experiment has a delayed overshoot, possibly as a result of a very weak AMOC in that GCM when CO2 is high. This study identifies aspects of overshoot behaviour that are robust across a multi-model and multi-scenario ensemble, and those that differ between experiments. These results could inform an assessment of the real-world AMOC response to decreasing CO2.