Auxiliary subunit regulation of high-voltage activated calcium channels expressed in mammalian cells

The effects of auxiliary calcium channel subunits on the expression and functional properties of high-voltage activated (HVA) calcium channels have been studied extensively in the Xenopus oocyte expression system, but are less completely characterized in a mammalian cellular environment. Here, we provide the first systematic analysis of the effects of calcium channel beta and alpha(2)-delta subunits on expression levels and biophysical properties of three different types (Ca(v)1.2, Ca(v)2.1 and Ca(v)2.3) of HVA calcium channels expressed in tsA-201 cells. Our data show that Ca(v)1.2 and Ca(v)2.3 channels yield significant barium current in the absence of any auxiliary subunits. Although calcium channel beta subunits were in principle capable of increasing whole cell conductance, this effect was dependent on the type of calcium channel alpha(1) subunit, and beta(3) subunits altogether failed to enhance current amplitude irrespective of channel subtype. Moreover, the alpha(2)-delta subunit alone is capable of increasing current amplitude of each channel type examined, and at least for members of the Ca(v)2 channel family, appears to act synergistically with beta subunits. In general agreement with previous studies, channel activation and inactivation gating was regulated both by beta and by alpha(2)-delta subunits. However, whereas pronounced regulation of inactivation characteristics was seen with the majority of the auxiliary subunits, effects on voltage dependence of activation were only small (< 5 mV). Overall, through a systematic approach, we have elucidated a previously underestimated role of the alpha(2)-delta(1) subunit with regard to current enhancement and kinetics. Moreover, the effects of each auxiliary subunit on whole cell conductance and channel gating appear to be specifically tailored to subsets of calcium channel subtypes

A Caenorhabditis elegans assay of seizure-like activity optimised for identifying antiepileptic drugs and their mechanisms of action

BACKGROUND: Epilepsy affects around 1% of people, but existing antiepileptic drugs (AEDs) only offer symptomatic relief and are ineffective in approximately 30% of patients. Hence, new AEDs are sorely needed. However, a major bottleneck is the low-throughput nature of early-stage AED screens in conventional rodent models. This process could potentially be expedited by using simpler invertebrate systems, such as the nematode Caenorhabditis elegans. NEW METHOD: Head-bobbing convulsions were previously reported to be inducible by pentylenetetrazol (PTZ) in C. elegans with loss-of-function mutations in unc-49, which encodes a GABAA receptor. Given that epilepsy-linked mutations in human GABAA receptors are well documented, this could represent a clinically-relevant system for early-stage AED screens. However, the original agar plate-based assay is unsuited to large-scale screening and has not been validated for identifying AEDs. Therefore, we established an alternative streamlined, higher-throughput approach whereby mutants were treated with PTZ and AEDs via liquid-based incubation. RESULTS: Convulsions induced within minutes of PTZ exposure in unc-49 mutants were strongly inhibited by the established AED ethosuximide. This protective activity was independent of ethosuximide's suggested target, the T-type calcium channel, as a null mutation in the worm cca-1 ortholog did not affect ethosuximide's anticonvulsant action. COMPARISON WITH EXISTING METHOD: Our streamlined assay is AED-validated, feasible for higher throughput compound screens, and can facilitate insights into AED mechanisms of action. CONCLUSIONS: Based on an epilepsy-associated genetic background, this C. elegans unc-49 model of seizure-like activity presents an ethical, higher throughput alternative to conventional rodent seizure models for initial AED screens

Knowlton Circles: A Later Neolithic and Early Bronze Age Ceremonial Complex and Its Environs—A Review

The larger henge monuments of Wessex have been the focus of detailed archaeological investigation for over a century and consequently their study has provided a greater, albeit fragmentary, understanding of later Neolithic society. For the most part such studies have continued to show that these great mega-structures of the third millennium B.C. persist as beacons of ceremonial functionality into which later societies invested much of themselves, intellectually, spiritually and unquestionably physically. While Stonehenge, Avebury and Durrington Walls continue to attract a great deal of attention with ongoing research, comparable sites in Dorset have been less well researched. Two campaigns of archaeological investigation undertaken in the Allen Valley of east Dorset by the author have focussed upon the complex of earthworks at Knowlton and additionally at one of three broadly contemporary barrow cemeteries located nearby. The findings from these investigations are beginning to shed more light on the possible origins and development of these important but weakly understood landscapes. This paper outlines some of the main findings from these investigations and posits a chronological framework for the integration of a group of monuments that formed both a ceremonial landscape and a geographical and spiritual home for communities that lasted for a thousand years

Taxonomy based on science is necessary for global conservation

Peer reviewe

Instabilities in liquid crystals

This thesis contains theoretical work dealing with the effects of magnetic and electric fields on samples of nematic, smectic A and smectic C liquid crystals. Some background material along with the continuum theory is introduced in Chapter 2. In Chapter 3 we consider the effect on the director within an infinite sample of nematic liquid crystal which is subjected to crossed electric and magnetic fields. In particular we examine the stability of the travelling waves which describe the director motion by considering the behaviour of the stable perturbations as time increases. The work of Chapter 4 examines a bounded sample of smectic A liquid crystal in two dimensions which is subjected to a magnetic field. It can be shown that layer undulations may occur after a critical threshold is reached. The effects of various forms of pressure are also studied and in the case when pressure is included it is shown that layer undulations will always occur regardless of the field strength. Chapter 5 extends the work of Chapter 4 to three dimensions. We consider the effect of adding a dynamical term and study the stability of these solutions. Also we examine how the layers undulate when the sample of smectic A liquid crystal is confined to various cylindrical geometries and subjected to a magnetic field and a uniform pressure. In Chapter 6 we consider a sample of smectic C liquid crystal which is subjected to an oscillatory electric field. The motion of the c-director is complicated and we examine the differences which occur for differing dielectric anisotropies. Finally, in Chapter 7 we study how a sample of smectic C liquid crystal behaves when it is subjected to a uniform shear flow within the smectic plane. We find travelling wave solutions for the behaviour of the c-director and adapt these solutions to incorporate the effects of an applied field. (author)Available from British Library Document Supply Centre-DSC:DXN045547 / BLDSC - British Library Document Supply CentreSIGLEGBUnited Kingdo

Theta method dynamics

Long-term solutions of the theta method applied to scalar nonlinear differential equations are studied in this paper. In the case where the equation has a stable steady state, lower bounds on the basin of non-oscillatory, monotonic attraction for the theta method are derived. Spurious period two solutions are then analysed. Under mild assumptions, precise results are obtained concerning the generic nature and stability of these solutions for small timesteps. Particular problem classes are studied, and direct connections are made between the existence and stability of period two solutions and the dynamics of the theta method. The analysis is extended to a wide class of semi-discretized partial differential equations. Numerical examples are given

Dissociation of phenotypic and functional endothelial progenitor cells in patients undergoing percutaneous coronary intervention

Objectives: Endothelial progenitor cells (EPCs) are circulating mononuclear cells with the capacity to mature into endothelial cells and contribute to vascular repair. We assessed the effect of local vascular injury during percutaneous coronary intervention (PCI) on circulating EPCs in patients with coronary artery disease. Design and setting: Prospective case-control study in a university teaching hospital. Patients: 54 patients undergoing elective coronary angiography. Interventions and main outcome measures: EPCs were quantified by flow cytometry (CD34+KDR+ phenotype) complemented by real-time polymerase chain reaction (PCR), and the colony forming unit (CFU-EC) functional assay, before and during the first 24 hours after diagnostic angiography (n = 27) or PCI (n = 27). Results: Coronary intervention, but not diagnostic angiography, resulted in an increase in blood neutrophil count (p&lt;0.001) and C-reactive protein concentrations (p = 0.001) in the absence of significant myocardial necrosis. Twenty-four hours after PCI, CFU-ECs increased threefold (median [IQR], 4.4 [1.3–13.8] vs 16.0 [2.1–35.0], p = 0.01), although circulating CD34+KDR+ cells (0.019% (SEM 0.004%) vs 0.016% (0.003%) of leucocytes, p = 0.62) and leucocyte CD34 mRNA (relative quantity 2.3 (0.5) vs 2.1 (0.4), p = 0.21) did not. There was no correlation between CFU-ECs and CD34+KDR+ cells. Conclusions: Local vascular injury following PCI results in a systemic inflammatory response and increases functional CFU-ECs. This increase was not associated with an early mobilisation of CD34+KDR+ cells, suggesting these cells are not the primary source of EPCs involved in the immediate response to vascular injury