Ratiometric and nonratiometric Ca2+ indicators for the assessment of intracellular free Ca2+ in a breast cancer cell line using a fluorescence microplate reader

Transporters of Ca2+ are potential drug targets and Ca2+ is a useful signal in the assessment of G-protein-coupled receptor activation. Assays involving the assessment of intracellular Ca2+ using microplate readers most often use Ca2+ indicators which do not exhibit a spectra shift on Ca2+ binding (e.g. fluo-3). Indicators that do exhibit a spectral shift upon Ca2+ binding (e.g. fura-2) offer potential advantages for the calibration of intracellular Ca2+ levels. However, experimental limitations may limit the use of ratiometric dyes in microplate readers capable of screening. In this study, we compared the assessment of intracellular Ca2+ in adherent breast cancer cells using ratiometric and nonratiometric Ca2+ indicators. Our results demonstrate that both fluo-3 and fura-2 detect ATP dose-dependent increases in intracellular Ca2+ in the MCF-7 breast cancer cell line and that some of the limitations in the use of fura-2 appear to be overcome by the use of glass bottom microplates. The calibrated intracellular Ca2+ levels derived using fura-2 are consistent with those from microscopy and cuvette-based studies. Fura-2 may be useful in microplate studies, where cell lines with different properties are compared or where screening treatments lead to differences in the number of cells or dye loading. (C) 2003 Elsevier B.V. All rights reserved.

Comparative adsorption of spherical argon and flexible n-butane in carbon slit pores - a GCMC computer simulation study

In this paper we investigate the difference between the adsorption of spherical molecule argon (at 87.3 K) and the flexible normal butane (at an equivalent temperature of 150 K) in carbon slit pores. These temperatures are equivalent in the sense that they have the same relative distances between their respective triple points and critical points. Higher equivalent temperatures are also studied (122.67 K for argon and 303 K for n-butane) to investigate the effects of temperature on the 2D-transition in adsorbed density. The Grand Canonical Monte Carlo simulation is used to study the adsorption of these two model adsorbates. Beside the longer computation times involved in the computation of n-butane adsorption, n-butane exhibits many interesting behaviors such as: (i) the onset of adsorption occurs sooner (in terms of relative pressure), (ii) the hysteresis for 2D- and 3D-transitions is larger, (iii) liquid-solid transition is not possible, (iv) 2D-transition occurs for n-butane at 150 K while it does not happen for argon except for pores that accommodate two layers of molecules, (v) the maximum pore density is about four times less than that of argon and (vi) the sieving pore width is slightly larger than that for argon. Finally another feature obtained from the Grand Canonical Monte Carlo (GCMC) simulation is the configurational arrangement of molecules in pores. For spherical argon, the arrangement is rather well structured, while for n-butane the arrangement depends very much on the pore size. (C) 2004 Elsevier B.V. All rights reserved.

Resolution enhancement by subtraction of confocal signals taken at different pinhole sizes

Subtractive imaging in confocal fluorescence light microscopy is based on the subtraction of a suitably weighted widefield image from a confocal image. An approximation to a widefield image can be obtained by detection with an opened confocal pinhole. The subtraction of images enhances the resolution in-plane as well as along the optic axis. Due to the linearity of the approach, the effect of subtractive imaging in Fourier-space corresponds to a reduction of low spatial frequency contributions leading to a relative enhancement of the high frequencies. Along the direction of the optic axis this also results in an improved sectioning. Image processing can achieve a similar effect. However, a 3D volume dataset must be acquired and processed, yielding a result essentially identical to subtractive imaging but superior in signal-to-noise ratio. The latter can be increased further with the technique of weighted averaging in Fourier-space. A comparison of 2D and 3D experimental data analysed with subtractive imaging, the equivalent Fourier-space processing of the confocal data only, and Fourier-space weighted averaging is presented. (C) 2003 Elsevier Ltd. All rights reserved.

Use of real-time three-dimensional echocardiography to measure left atrial volume: Comparison with other echocardiographic techniques

Objectives: Left atrial (LA) volume (LAV) is a prognostically important biomarker for diastolic dysfunction, but its reproducibility on repeated testing is not well defined. LA assessment with 3-dimensional. (3D) echocardiography (3DE) has been validated against magnetic resonance imaging, and we sought to assess whether this was superior to existing measurements for sequential echocardiographic follow-up. Methods: Patients (n = 100; 81 men; age 56 +/- 14 years) presenting for LA evaluation were studied with M-mode (MM) echocardiography, 2-dimensional (2D) echocardiography, and 3DE. Test-retest variation was performed by a complete restudy by a separate sonographer within 1 hour without alteration of hemodynamics or therapy. In all, 20 patients were studied for interobserver and intraobserver variation. LAVs were calculated by using M-mode diameter and planimetered atrial area in the apical. 4-chamber view to calculate an assumed sphere, as were prolate ellipsoid, Simpson's biplane, and biplane area-length methods. All were compared with 3DE. Results: The average LAV was 72 +/- 27 mL by 3DE. There was significant underestimation of LAV by M-mode (35 +/- 20 mL, r = 0.66, P < .01). The 3DE and various 2D echocardiographic techniques were well correlated: LA planimetry (85 +/- 38 mL, r = 0.77, P < .01), prolate ellipsoid (73 +/- 36 mL, r = 0.73, P = .04), area-length (64 +/- 30 mL, r = 0.74, P < .01), and Simpson's biplane (69 +/- 31 mL, r = 0.78, P = .06). Test-retest variation for 3DE was most favorable (r = 0.98, P < .01), with the prolate ellipsoid method showing most variation. Interobserver agreement between measurements was best for 3DE (r = 0.99, P < .01), with M-mode the worst (r = 0.89, P < .01). Intraobserver results were similar to interobserver, the best correlation for 3DE (r = 0.99, P < .01), with LA planimetry the worst (r = 0.91, P < .01). Conclusions. The 2D measurements correlate closely with 3DE. Follow-up assessment in daily practice appears feasible and reliable with both 2D and 3D approaches.

A theoretical comparison of two optimization methods for radiofrequency drive schemes in high frequency MRI resonators

In this paper, numerical simulations are used in an attempt to find optimal Source profiles for high frequency radiofrequency (RF) volume coils. Biologically loaded, shielded/unshielded circular and elliptical birdcage coils operating at 170 MHz, 300 MHz and 470 MHz are modelled using the FDTD method for both 2D and 3D cases. Taking advantage of the fact that some aspects of the electromagnetic system are linear, two approaches have been proposed for the determination of the drives for individual elements in the RF resonator. The first method is an iterative optimization technique with a kernel for the evaluation of RF fields inside an imaging plane of a human head model using pre-characterized sensitivity profiles of the individual rungs of a resonator; the second method is a regularization-based technique. In the second approach, a sensitivity matrix is explicitly constructed and a regularization procedure is employed to solve the ill-posed problem. Test simulations show that both methods can improve the B-1-field homogeneity in both focused and non-focused scenarios. While the regularization-based method is more efficient, the first optimization method is more flexible as it can take into account other issues such as controlling SAR or reshaping the resonator structures. It is hoped that these schemes and their extensions will be useful for the determination of multi-element RF drives in a variety of applications.

An anisotropic viscous representation of Mohr-Coulomb failure for use is modelling coupled mantle-continent dynamics

In mantle convection models it has become common to make use of a modified (pressure sensitive, Boussinesq) von Mises yield criterion to limit the maximum stress the lithosphere can support. This approach allows the viscous, cool thermal boundary layer to deform in a relatively plate-like mode even in a fully Eulerian representation. In large-scale models with embedded continental crust where the mobile boundary layer represents the oceanic lithosphere, the von Mises yield criterion for the oceans ensures that the continents experience a realistic broad-scale stress regime. In detailed models of crustal deformation it is, however, more appropriate to choose a Mohr-Coulomb yield criterion based upon the idea that frictional slip occurs on whichever one of many randomly oriented planes happens to be favorably oriented with respect to the stress field. As coupled crust/mantle models become more sophisticated it is important to be able to use whichever failure model is appropriate to a given part of the system. We have therefore developed a way to represent Mohr-Coulomb failure within a code which is suited to mantle convection problems coupled to large-scale crustal deformation. Our approach uses an orthotropic viscous rheology (a different viscosity for pure shear to that for simple shear) to define a prefered plane for slip to occur given the local stress field. The simple-shear viscosity and the deformation can then be iterated to ensure that the yield criterion is always satisfied. We again assume the Boussinesq approximation - neglecting any effect of dilatancy on the stress field. An additional criterion is required to ensure that deformation occurs along the plane aligned with maximum shear strain-rate rather than the perpendicular plane which is formally equivalent in any symmetric formulation. It is also important to allow strain-weakening of the material. The material should remember both the accumulated failure history and the direction of failure. We have included this capacity in a Lagrangian-Integration-point finite element code and will show a number of examples of extension and compression of a crustal block with a Mohr-Coulomb failure criterion, and comparisons between mantle convection models using the von Mises versus the Mohr-Coulomb yield criteria. The formulation itself is general and applies to 2D and 3D problems, although it is somewhat more complicated to identify the slip plane in 3D.

Mental health nursing standards and practice indicators for Australia: a review of current literature

There are many changes and challenges facing the mental health care professional working in Australia in the 21st Century. Given the significance of their number and the considerable extent to which care is delivered by them, mental health nurses in particular must be at the forefront of the movement to enhance and improve mental health care. Mental health nurses in Australia must not only keep up with the changes, we should be setting the pace for others across the profession worldwide. The increasingly complex field of mental health nursing demands nurses who are not only equipped to face the challenges but are confident in doing so. Definitive guidelines for practice, clear expectations regarding outcomes and specific means by which to evaluate both practice and outcomes are vital. Strengthening the role and vision of mental health nursing so that there is clarity about both and highlighting core values by which to perform will enable us to become focused on our future and what we can expect to both give to and receive from our chosen profession and how we can, and do, contribute to mental health care. The role of the mental health nurse is undergoing expansion and there are new hurdles to overcome along with the new benefits this brings. To support this, nationally adopted, formalised standards of practice and means by which to measure these, i.e., practice indicators formerly known as clinical indicators, are required. It is important to have national standards and practice indicators because of the variances in the provision of mental health across Australia – different legislation regarding mental health policies and processes, different nursing registration bodies and Nursing Councils, for example – which create additional barriers to cohesion and uniformity. Improvements in the practice of mental health nursing lead to benefits for consumer outcomes as well as the overall quality of mental health care available in Australia. The emphasis on rights-based care, particularly consumer and carer rights, demands evidence-based, up-to-date mental health care delivered by competent, capable professionals. Documented expectations for performance by nurses will provide all involved with yardsticks by which to evaluate outcomes. Flowing on from these benefits are advances in mental health care generally and enhancements to Australia’s reputation and position within the health care arena throughout the world. Currently, the ‘Standards for Practice’ published by the Australian New Zealand College of Mental Health Nurses (ANZCMHN) in 1995 and the practice indicators developed by Skews et al. (2000) provide a less formal guide for mental health nurses working in Australia. While these earlier standards and practice indicators have played some role in supporting mental health nurses they have not been nationally or enthusiastically adopted and there are a multitude of reasons for this. This report reviews the current literature available on practice indicators and standards for practice and describes an evidence-based rationale as to why a review and renewal of these is required and why it is important, not just for mental health nurses but to the field of mental health in general. The term ‘practice indicator’ is used, except where a quotation utilises ‘clinical indicator’, to more accurately reflect the broad spectrum of nursing roles, i.e. not all mental health nursing work involves a clinical role.

Urban and rural suicide differentials in migrants and the Australian-born, New South Wales, Australia 1985-1994

We estimated risk of suicide in adults in New South Wales (NSW) by sex, country of birth and rural/urban residence, after adjusting for age; we also examined youth suicide (age 15-24 years). The study population was the entire population of NSW, Australia, aged greater than or equal to 15 years during the period 1985-1994. Poisson regression was used to examine the relationship between predictor variables and the risk of suicide, with the focus on migrant status and area of residence. A significantly higher risk of suicide was found in male migrants from Northern Europe and Eastern Europe/former USSR, compared to Australian-born males; a significantly lower suicide risk occurred in males from Southern Europe, the Middle East and Asia. In female migrants, those from UK/Eire, Northern Europe, Eastern Europe/former USSR and New Zealand exhibited a significantly higher risk of suicide compared to Australian-born females. A significantly lower risk of suicide occurred in females from the Middle East. Male migrants overall were at significantly lower risk of suicide than the Australian-born, while female migrants overall had a significantly higher risk of suicide than Australian-born females. Among migrant males overall, the rural-urban suicide risk differential was significantly higher for those living in non-metropolitan areas (RR = 1.9; 95% CI: 1.7-2.1). Suicide risk was significantly higher in non-metropolitan male immigrants from the UK/Eire (RR = 1.4; 95% CI: 1.1-1.7), Southern Europe (RR = 1.7; 95% CI: 1.2-2.4), Northern/Western Europe (1.5; 95% CI: 1.2-1.9), the Middle East (RR = 3.8; 95% CI: 1.9-7.8), New :Zealand (RR = 1.4; 95% CI: 1.0-1.8) and 'other' (RR = 2.6; 95% CI: 1.9-3.5), when compared to their urban counterparts. There was no statistically significant difference in suicide risk between rural and urban Australian-born males. For female suicide, significantly lower risk was found in female immigrants living in non-metropolitan areas who were from Northern/Western Europe (RR = 0.7; 95% CI: 0.4-0.96), as well as the Australian-born (RR = 0.7; 95% CI: 0.6-0.8), when compared to their urban counterparts. The non-metropolitan/metropolitan relative risk for suicide in female migrants overall was not significantly different from one. Among male youth there was a significantly higher suicide risk in non-metropolitan areas, with a relative risk estimate of 1.4 for Australian-born youth (95% CI: 1.2-1.5) and 1.7 for migrant youth (95% CI: 1.2-2.4), when compared with metropolitan counterparts. We conclude that suicide among migrant males living in non-metropolitan areas accounts for most of the excess of male suicide in rural NSW, and the significantly lower risk of suicide for non-metropolitan Australian-born women does not apply to migrant women. (C) 1999 Published by Elsevier Science Ltd. All rights reserved.

Differences in physical activity between black and white girls living in rural and urban areas

This study examined the relationship of race and rural/urban setting to physical, behavioral, psychosocial, and environmental factors associated with physical activity. Subjects included 1,668 eighth-grade girls from 31 middle schools: 933 from urban settings, and 735 from rural settings. Forty-six percent of urban girls and 59% of rural girls were Black. One-way and two-way ANOVAs with school as a covariate were used to analyze the data. Results indicated that most differences were associated with race rather than setting. Black girls were less active than White girls, reporting significantly fewer 30-minute blocks of both vigorous and moderate-to-vigorous physical activity. Black girls also spent more time watching television, and had higher BMIs and greater prevalence of overweight than White girls. However, enjoyment of physical education and family involvement in physical activity were greater among Black girls titan White girls. Rural White girls and urban Black girls had more favorable attitudes toward physical activity. Access to sports equipment, perceived safety of neighborhood, and physical activity self-efficacy were higher in White girls than Black girls.

Motivations and perceived influences on rural and urban general practitioners when prescribing conventional non-steroidal anti-inflammatory drugs or COX-2 inhibitors

Background and objective: Prescribers in rural and remote locations perceive that there are different influences on their prescribing compared with those experienced by urban prescribers. The aim of this study was to compare the motivations and perceived influences on general practitioners (GPs) when prescribing COX-2 inhibitors rather than conventional non-steroidal anti-inflammatory drugs (NSAIDs) between rural and urban-based GPs in Queensland, Australia. Methods: A questionnaire was administered to two geographically distinct groups of GPs, one urban (n = 67) and one rural (n = 67), investigating the reasons that the GP would prescribe a COX-2 inhibitor rather than a conventional NSAID or vice versa and also focusing on patients requesting a prescription for a COX-2 inhibitor. Results and discussion: A 51% response rate (n = 68) was achieved. The difference between the rural and the urban GPs was that the urban GPs were more likely to perceive that they were influenced to prescribe COX-2 inhibitors by their patients' knowledge of these new (at the time) drugs. GPs in both the rural and urban areas perceived the COX-2 selective inhibitors to be safer than conventional NSAIDs, and that there was little difference in terms of efficacy between the two drug classes. However, GPs from both of the study areas stated that conventional NSAIDs were preferred over COX-2 selective inhibitors, primarily due to their expense, if their patients were not at risk for developing a GI bleed. Conclusion: The motivations and perceived influences to prescribe a COX-2 inhibitor in rural and in urban areas of Queensland, Australia were very similar. Almost all surveyed GPs in rural and urban areas had patients request a prescription, or enquire about the COX-2 inhibitors. Urban GPs were more likely to feel pressured to prescribe a COX-2 inhibitor than their rural counterparts, agreeing with other research which found that patient pressure to prescribe appears to be greater in urban general practice.