A polymorphism in the norepinephrine transporter gene is associated with affective and cardiovascular disease through a microRNA mechanism.

Norepinephrine released from sympathetic nerves is removed from the neuroeffector junction via the action of the norepinephrine transporter (NET). NET impairment is evident in several clinically important conditions including major depressive disorder (MDD), panic disorder (PD), essential hypertension and the postural orthostatic tachycardia syndrome (POTS). We aimed to determine whether a single nucleotide polymorphism (SNP) in the 3' untranslated region (UTR) of the NET gene is associated with NET impairment and to elucidate the mechanisms involved. The analyses were carried out in two cohorts of European ancestry, which included healthy controls and MDD, PD, hypertensive and POTS patients. Compared with controls, cases had significantly higher prevalence of the T allele of rs7194256 (C/T), arterial norepinephrine, depression and anxiety scores, larger left ventricular mass index, higher systolic and diastolic blood pressures, and heart rate. Bioinformatic analysis identified that the microRNA miR-19a-3p could bind preferentially to the sequence created by the presence of the T allele. This was supported by results of luciferase assays. Compared with controls, cases had significantly lower circulating miR-19a-3p, which was associated with pathways related to blood pressure and regulation of neurotransmission. In vitro norepinephrine downregulated miR-19a-3p. In conclusion, the T allele of the rs7194256 SNP in the 3'UTR of the NET gene is more prevalent in diseases where NET impairment is evident. This might be explained by the creation of a binding site for the microRNA miR-19a-3p. A defect in NET function may potentiate the sympathetic neurochemical signal, predisposing individuals with affective diseases to increased risk of cardiovascular disease development.

Cryofixation rapidly preserves cytoskeletal arrays of leaf epidermal cells revealing microtubule co-alignments between neighbouring cells and adjacent actin and microtubule bundles in the cortex.

Accurate preservation of microtubule and actin microfilament arrays is crucial for investigating their roles in plant cell development. Aldehyde fixatives such as paraformaldehyde or glutaraldehyde preserve cortical microtubule arrays but, unless actin microfilaments are stabilized with drugs such as m-maleimidobenzoyl N-hydroxysuccinimide ester (MBS), ethylene glycol bis[sulfosuccinimidylsuccinate] (sulfo-EGS) or phalloidin, their arrays are often poorly preserved. Cryofixation, used primarily for electron microscopy, preserves actin microfilaments well but is used rarely to fix plant cells for optical microscopy. We developed a novel whole-mount cryofixation method to preserve microtubule and microfilament arrays within Tradescantia virginiana leaf epidermal cells for investigation using confocal microscopy. Cortical microtubule arrays were often oriented in different directions on the internal and external faces of the epidermal cells. A number of arrays were aligned in several directions, parallel to microtubules of neighbouring cells. Actin microfilaments were particularly well preserved possibly due to the speed with which they were immobilized. No transverse cortical microfilament arrays were observed. On occasion, we observed co-aligned microfilament and microtubule bundles lying adjacent to the plasma membrane and positioned side by side suggesting a potential direct interaction between the cytoskeletal filaments at these locations. Cryofixation is therefore a valuable tool to investigate the interactions between cytoskeletal arrays in plant cells using confocal microscopy.

Towards correlative imaging of plant cortical microtubule arrays: combining ultrastructure with real-time microtubule dynamics.

There are a variety of microscope technologies available to image plant cortical microtubule arrays. These can be applied specifically to investigate direct questions relating to array function, ultrastructure or dynamics. Immunocytochemistry combined with confocal laser scanning microscopy provides low resolution "snapshots" of cortical microtubule arrays at the time of fixation whereas live cell imaging of fluorescent fusion proteins highlights the dynamic characteristics of the arrays. High-resolution scanning electron microscopy provides surface detail about the individual microtubules that form cortical microtubule arrays and can also resolve cellulose microfibrils that form the innermost layer of the cell wall. Transmission electron microscopy of the arrays in cross section can be used to examine links between microtubules and the plasma membrane and, combined with electron tomography, has the potential to provide a complete picture of how individual microtubules are spatially organized within the cortical cytoplasm. Combining these high-resolution imaging techniques with the expression of fluorescent cytoskeletal fusion proteins in live cells using correlative microscopy procedures will usher in an radical change in our understanding of the molecular dynamics that underpin the organization and function of the cytoskeleton.

Correlative fluorescence and transmission electron microscopy: an elegant tool to study the actin cytoskeleton of whole-mount (breast) cancer cells.

Elucidating the structure and dynamics of lamellipodia and filopodia in response to different stimuli is a topic of continuing interest in cancer cells as these structures may be attractive targets for therapeutic purposes. Interestingly, a close functional relationship between these actin-rich protrusions and specialized membrane domains has been recently demonstrated. The aim of this study was therefore to investigate the fine organization of these actin-rich structures and examine how they structurally may relate to detergent-resistant membrane (DRM) domains in the MTLn3 EGF/serum starvation model. For this reason, we designed a straightforward and alternative method to study cytoskeleton arrays and their associated structures by means of correlative fluorescence (/laser)- and electron microscopy (CFEM). CFEM on whole mounted breast cancer cells revealed that a lamellipodium is composed of an intricate filamentous actin web organized in various patterns after different treatments. Both actin dots and DRM's were resolved, and were closely interconnected with the surrounding cytoskeleton. Long actin filaments were repeatedly observed extending beyond the leading edge and their density and length varied after different treatments. Furthermore, CFEM also allowed us to demonstrate the close structural association of DRMs with the cytoskeleton in general and the filamentous/dot-like structural complexes in particular, suggesting that they are all functionally linked and consequently may regulate the cell's fingertip dynamics. Finally, electron tomographic modelling on the same CFEM samples confirmed that these extensions are clearly embedded within the cytoskeletal matrix of the lamellipodium.

Force appropriation of nonlinear structures.

Nonlinear normal modes (NNMs) are widely used as a tool for developing mathematical models of nonlinear structures and understanding their dynamics. NNMs can be identified experimentally through a phase quadrature condition between the system response and the applied excitation. This paper demonstrates that this commonly used quadrature condition can give results that are significantly different from the true NNM, in particular, when the excitation applied to the system is limited to one input force, as is frequently used in practice. The system studied is a clamped-clamped cross-beam with two closely spaced modes. This paper shows that the regions where the quadrature condition is (in)accurate can be qualitatively captured by analysing transfer of energy between the modes of the system, leading to a discussion of the appropriate number of input forces and their locations across the structure.

Identifying the significance of nonlinear normal modes.

Nonlinear normal modes (NNMs) are widely used as a tool for understanding the forced responses of nonlinear systems. However, the contemporary definition of an NNM also encompasses a large number of dynamic behaviours which are not observed when a system is forced and damped. As such, only a few NNMs are required to understand the forced dynamics. This paper firstly demonstrates the complexity that may arise from the NNMs of a simple nonlinear system-highlighting the need for a method for identifying the significance of NNMs. An analytical investigation is used, alongside energy arguments, to develop an understanding of the mechanisms that relate the NNMs to the forced responses. This provides insight into which NNMs are pertinent to understanding the forced dynamics, and which may be disregarded. The NNMs are compared with simulated forced responses to verify these findings.

Doctoral dissertations on hypnosis: 1980-1989.

This article extends Clark, Hungerford, and Reilley's (1984) review of dissertations accepted by American universities from 1923 to 1980 by examining dissertations of the 1980s. A table shows which areas of research received the most attention in each of the last 3 decades. There was an increase in the percentage of psychology and educational psychology researchers using hypnosis in dissertations.

Auditory P300 studies in schizophrenic subjects and their first degree relatives.

Nineteen subjects with schizophrenia, 6 subjects with related disorders (schizophrenic spectrum disorders (S.S.D.)), and 20 unaffected first degree relatives from a sample of schizophrenic pedigrees, together with 35 normal control subjects, had auditory P300 evoked responses measured, using the "odd-ball" paradigm for stimulation. Bipolar recording on the midline (CZOZ) and two contralateral sites (CZ-mastoid) was carried out. It was found that approximately 40% of schizophrenics/S.S.D.'s had abnormal P300 responses. Abnormalities were seen in latency, RMS response voltage and in the left side response--right side response cross correlation coefficient. Schizophrenics/S.S.D.s showed responses which were topographically different than those of normal controls. Significant left-side/right-side response voltage asymmetry was not observed. In our study, only 10% of unaffected relatives of schizophrenics/S.S.D.'s showed abnormal P300 responses.

Systematic experimental exploration of bifurcations with noninvasive control.

We present a general method for systematically investigating the dynamics and bifurcations of a physical nonlinear experiment. In particular, we show how the odd-number limitation inherent in popular noninvasive control schemes, such as (Pyragas) time-delayed or washout-filtered feedback control, can be overcome for tracking equilibria or forced periodic orbits in experiments. To demonstrate the use of our noninvasive control, we trace out experimentally the resonance surface of a periodically forced mechanical nonlinear oscillator near the onset of instability, around two saddle-node bifurcations (folds) and a cusp bifurcation.

Correlative microscopy: providing new understanding in the biomedical and plant sciences.

Correlative microscopy is the application of two or more distinct microscopy techniques to the same region of a sample, generating complementary morphological, structural and chemical information that exceeds what is possible with any single technique. As a variety of complementary microscopy approaches rather than a specific type of instrument, correlative microscopy has blossomed in recent years as researchers have recognised that it is particularly suited to address the intricate questions of the modern biological sciences. Specialised technical developments in sample preparation, imaging methods, visualisation and data analysis have also accelerated the uptake of correlative approaches. In light of these advances, this critical review takes the reader on a journey through recent developments in, and applications of, correlative microscopy, examining its impact in biomedical research and in the field of plant science. This twin emphasis gives a unique perspective into use of correlative microscopy in fields that often advance independently, and highlights the lessons that can be learned from both fields for the future of this important area of research.

Membrane-wall attachments in plasmolysed plant cells.

Field emission scanning electron microscopy of plasmolysed Tradescantia virginiana leaf epidermal cells gave novel insights into the three-dimensional architecture of Hechtian strands, Hechtian reticulum, and the inner surface of the cell wall without the need for extraction. At high magnification, we observed fibres that pin the plasma membrane to the cell wall after plasmolysis. Treatment with cellulase caused these connecting fibres to be lost and the pinned out plasma membrane of the Hechtian reticulum to disintegrate into vesicles with diameters of 100-250 nm. This suggests that the fibres may be cellulose. After 4 h of plasmolysis, a fibrous meshwork that labelled with anti-callose antibodies was observed within the space between the plasmolysed protoplast and the cell wall by field emission scanning electron microscopy. Interestingly, macerase-pectinase treatment resulted in the loss of this meshwork, suggesting that it was stabilised by pectins. We suggest that cellulose microfibrils extending from strands of the Hechtian reticulum and entwining into the cell wall matrix act as anchors for the plasma membrane as it moves away from the wall during plasmolysis.