Calmodulin Activation by Calcium Transients in the Postsynaptic Density of Dendritic Spines

The entry of calcium into dendritic spines can trigger a sequence of biochemical reactions that begins with the activation of calmodulin (CaM) and ends with long-term changes to synaptic strengths. The degree of activation of CaM can depend on highly local elevations in the concentration of calcium and the duration of transient increases in calcium concentration. Accurate measurement of these local changes in calcium is difficult because the spaces are so small and the numbers of molecules are so low. We have therefore developed a Monte Carlo model of intracellular calcium dynamics within the spine that included calcium binding proteins, calcium transporters and ion channels activated by voltage and glutamate binding. The model reproduced optical recordings using calcium indicator dyes and showed that without the dye the free intracellular calcium concentration transient was much higher than predicted from the fluorescent signal. Excitatory postsynaptic potentials induced large, long-lasting calcium gradients across the postsynaptic density, which activated CaM. When glutamate was released at the synapse 10 ms before an action potential occurred, simulating activity patterns that strengthen hippocampal synapses, the calcium gradient and activation of CaM in the postsynaptic density were much greater than when the order was reversed, a condition that decreases synaptic strengths, suggesting a possible mechanism underlying the induction of long-term changes in synaptic strength. The spatial and temporal mechanisms for selectivity in CaM activation demonstrated here could be used in other signaling pathways

Two-Photon Imaging of Calcium in Virally Transfected Striate Cortical Neurons of Behaving Monkey

Two-photon scanning microscopy has advanced our understanding of neural signaling in non-mammalian species and mammals. Various developments are needed to perform two-photon scanning microscopy over prolonged periods in non-human primates performing a behavioral task. In striate cortex in two macaque monkeys, cortical neurons were transfected with a genetically encoded fluorescent calcium sensor, memTNXL, using AAV1 as a viral vector. By constructing an extremely rigid and stable apparatus holding both the two-photon scanning microscope and the monkey's head, single neurons were imaged at high magnification for prolonged periods with minimal motion artifacts for up to ten months. Structural images of single neurons were obtained at high magnification. Changes in calcium during visual stimulation were measured as the monkeys performed a fixation task. Overall, functional responses and orientation tuning curves were obtained in 18.8% of the 234 labeled and imaged neurons. This demonstrated that the two-photon scanning microscopy can be successfully obtained in behaving primates

Why Are Outcomes Different for Registry Patients Enrolled Prospectively and Retrospectively? Insights from the Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF).

Background: Retrospective and prospective observational studies are designed to reflect real-world evidence on clinical practice, but can yield conflicting results. The GARFIELD-AF Registry includes both methods of enrolment and allows analysis of differences in patient characteristics and outcomes that may result. Methods and Results: Patients with atrial fibrillation (AF) and ≥1 risk factor for stroke at diagnosis of AF were recruited either retrospectively (n = 5069) or prospectively (n = 5501) from 19 countries and then followed prospectively. The retrospectively enrolled cohort comprised patients with established AF (for a least 6, and up to 24 months before enrolment), who were identified retrospectively (and baseline and partial follow-up data were collected from the emedical records) and then followed prospectively between 0-18 months (such that the total time of follow-up was 24 months; data collection Dec-2009 and Oct-2010). In the prospectively enrolled cohort, patients with newly diagnosed AF (≤6 weeks after diagnosis) were recruited between Mar-2010 and Oct-2011 and were followed for 24 months after enrolment. Differences between the cohorts were observed in clinical characteristics, including type of AF, stroke prevention strategies, and event rates. More patients in the retrospectively identified cohort received vitamin K antagonists (62.1% vs. 53.2%) and fewer received non-vitamin K oral anticoagulants (1.8% vs . 4.2%). All-cause mortality rates per 100 person-years during the prospective follow-up (starting the first study visit up to 1 year) were significantly lower in the retrospective than prospectively identified cohort (3.04 [95% CI 2.51 to 3.67] vs . 4.05 [95% CI 3.53 to 4.63]; p = 0.016). Conclusions: Interpretations of data from registries that aim to evaluate the characteristics and outcomes of patients with AF must take account of differences in registry design and the impact of recall bias and survivorship bias that is incurred with retrospective enrolment. Clinical Trial Registration: - URL: http://www.clinicaltrials.gov . Unique identifier for GARFIELD-AF (NCT01090362)

Improved risk stratification of patients with atrial fibrillation: an integrated GARFIELD-AF tool for the prediction of mortality, stroke and bleed in patients with and without anticoagulation.

OBJECTIVES: To provide an accurate, web-based tool for stratifying patients with atrial fibrillation to facilitate decisions on the potential benefits/risks of anticoagulation, based on mortality, stroke and bleeding risks. DESIGN: The new tool was developed, using stepwise regression, for all and then applied to lower risk patients. C-statistics were compared with CHA2DS2-VASc using 30-fold cross-validation to control for overfitting. External validation was undertaken in an independent dataset, Outcome Registry for Better Informed Treatment of Atrial Fibrillation (ORBIT-AF). PARTICIPANTS: Data from 39 898 patients enrolled in the prospective GARFIELD-AF registry provided the basis for deriving and validating an integrated risk tool to predict stroke risk, mortality and bleeding risk. RESULTS: The discriminatory value of the GARFIELD-AF risk model was superior to CHA2DS2-VASc for patients with or without anticoagulation. C-statistics (95% CI) for all-cause mortality, ischaemic stroke/systemic embolism and haemorrhagic stroke/major bleeding (treated patients) were: 0.77 (0.76 to 0.78), 0.69 (0.67 to 0.71) and 0.66 (0.62 to 0.69), respectively, for the GARFIELD-AF risk models, and 0.66 (0.64-0.67), 0.64 (0.61-0.66) and 0.64 (0.61-0.68), respectively, for CHA2DS2-VASc (or HAS-BLED for bleeding). In very low to low risk patients (CHA2DS2-VASc 0 or 1 (men) and 1 or 2 (women)), the CHA2DS2-VASc and HAS-BLED (for bleeding) scores offered weak discriminatory value for mortality, stroke/systemic embolism and major bleeding. C-statistics for the GARFIELD-AF risk tool were 0.69 (0.64 to 0.75), 0.65 (0.56 to 0.73) and 0.60 (0.47 to 0.73) for each end point, respectively, versus 0.50 (0.45 to 0.55), 0.59 (0.50 to 0.67) and 0.55 (0.53 to 0.56) for CHA2DS2-VASc (or HAS-BLED for bleeding). Upon validation in the ORBIT-AF population, C-statistics showed that the GARFIELD-AF risk tool was effective for predicting 1-year all-cause mortality using the full and simplified model for all-cause mortality: C-statistics 0.75 (0.73 to 0.77) and 0.75 (0.73 to 0.77), respectively, and for predicting for any stroke or systemic embolism over 1 year, C-statistics 0.68 (0.62 to 0.74). CONCLUSIONS: Performance of the GARFIELD-AF risk tool was superior to CHA2DS2-VASc in predicting stroke and mortality and superior to HAS-BLED for bleeding, overall and in lower risk patients. The GARFIELD-AF tool has the potential for incorporation in routine electronic systems, and for the first time, permits simultaneous evaluation of ischaemic stroke, mortality and bleeding risks. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique identifier for GARFIELD-AF (NCT01090362) and for ORBIT-AF (NCT01165710)

Two-year outcomes of patients with newly diagnosed atrial fibrillation: results from GARFIELD-AF.

AIMS: The relationship between outcomes and time after diagnosis for patients with non-valvular atrial fibrillation (NVAF) is poorly defined, especially beyond the first year. METHODS AND RESULTS: GARFIELD-AF is an ongoing, global observational study of adults with newly diagnosed NVAF. Two-year outcomes of 17 162 patients prospectively enrolled in GARFIELD-AF were analysed in light of baseline characteristics, risk profiles for stroke/systemic embolism (SE), and antithrombotic therapy. The mean (standard deviation) age was 69.8 (11.4) years, 43.8% were women, and the mean CHA2DS2-VASc score was 3.3 (1.6); 60.8% of patients were prescribed anticoagulant therapy with/without antiplatelet (AP) therapy, 27.4% AP monotherapy, and 11.8% no antithrombotic therapy. At 2-year follow-up, all-cause mortality, stroke/SE, and major bleeding had occurred at a rate (95% confidence interval) of 3.83 (3.62; 4.05), 1.25 (1.13; 1.38), and 0.70 (0.62; 0.81) per 100 person-years, respectively. Rates for all three major events were highest during the first 4 months. Congestive heart failure, acute coronary syndromes, sudden/unwitnessed death, malignancy, respiratory failure, and infection/sepsis accounted for 65% of all known causes of death and strokes for <10%. Anticoagulant treatment was associated with a 35% lower risk of death. CONCLUSION: The most frequent of the three major outcome measures was death, whose most common causes are not known to be significantly influenced by anticoagulation. This suggests that a more comprehensive approach to the management of NVAF may be needed to improve outcome. This could include, in addition to anticoagulation, interventions targeting modifiable, cause-specific risk factors for death. CLINICAL TRIAL REGISTRATION: http://www.clinicaltrials.gov. Unique identifier: NCT01090362

Risk profiles and one-year outcomes of patients with newly diagnosed atrial fibrillation in India: Insights from the GARFIELD-AF Registry.

BACKGROUND: The Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF) is an ongoing prospective noninterventional registry, which is providing important information on the baseline characteristics, treatment patterns, and 1-year outcomes in patients with newly diagnosed non-valvular atrial fibrillation (NVAF). This report describes data from Indian patients recruited in this registry. METHODS AND RESULTS: A total of 52,014 patients with newly diagnosed AF were enrolled globally; of these, 1388 patients were recruited from 26 sites within India (2012-2016). In India, the mean age was 65.8 years at diagnosis of NVAF. Hypertension was the most prevalent risk factor for AF, present in 68.5% of patients from India and in 76.3% of patients globally (P < 0.001). Diabetes and coronary artery disease (CAD) were prevalent in 36.2% and 28.1% of patients as compared with global prevalence of 22.2% and 21.6%, respectively (P < 0.001 for both). Antiplatelet therapy was the most common antithrombotic treatment in India. With increasing stroke risk, however, patients were more likely to receive oral anticoagulant therapy [mainly vitamin K antagonist (VKA)], but average international normalized ratio (INR) was lower among Indian patients [median INR value 1.6 (interquartile range {IQR}: 1.3-2.3) versus 2.3 (IQR 1.8-2.8) (P < 0.001)]. Compared with other countries, patients from India had markedly higher rates of all-cause mortality [7.68 per 100 person-years (95% confidence interval 6.32-9.35) vs 4.34 (4.16-4.53), P < 0.0001], while rates of stroke/systemic embolism and major bleeding were lower after 1 year of follow-up. CONCLUSION: Compared to previously published registries from India, the GARFIELD-AF registry describes clinical profiles and outcomes in Indian patients with AF of a different etiology. The registry data show that compared to the rest of the world, Indian AF patients are younger in age and have more diabetes and CAD. Patients with a higher stroke risk are more likely to receive anticoagulation therapy with VKA but are underdosed compared with the global average in the GARFIELD-AF. CLINICAL TRIAL REGISTRATION-URL: http://www.clinicaltrials.gov. Unique identifier: NCT01090362

Application of electro-membrane processes to desalination of solutions containing organic compounds

Przedstawiono możliwości procesów elektromembranowych w odsalaniu wodnych roztworów zawierających substancje organiczne. Dyskusję poprzedzono syntetycznym opisem konwencjonalnej elektrodializy, elektrodializy odwracalnej i elektro-dejonizacji. Podkreślono, że zagadnienie odsalania roztworów substancji organicznych jest ważne nie tylko w technologii biochemicznej, ale też w oczyszczaniu wód technologicznych i ścieków przemysłowych. Odsalanie i zatężanie ścieków przemysłowych w układzie mikrofiltracja–elektrodializa pozwala na odzyskanie wody i wartościowych substancji, dając możliwość tworzenia zamkniętych obiegów wody. Wykazano, że usuwanie soli z roztworów zawierających obojętne makrocząsteczki organiczne może być w prosty sposób prowadzone za pomocą konwencjonalnej elektrodializy. Obecność substancji organicznych o jonowym charakterze powoduje intensywne blokowanie membran (fouling) i/lub przenikanie składników organicznych do strumienia koncentratu. Stwierdzono, że w celu wyeliminowania tych problemów należy stosować elektrodializę odwracalną i jonoselektywne membrany jonowymienne.The potential of electro-membrane processes in salt separation from aqueous solutions containing organic substances was discussed. The discussion was preceded by brief description of conventional electrodialysis, electrodialysis reversal and electrodeionization. It was emphasized that desalination of organic substance solutions was essential not only for biochemical technology, but also for industrial water and wastewater treatment. Desalination and concentration of industrial effluents by microfiltration–electrodialysis system enabled reuse of water and valuable substances, thus providing the possibility of creating water closed-loop systems. It was demonstrated that salt removal from solutions containing neutral organic macroparticles might be a simple procedure when conducted with use of conventional electrodialysis. In contrast, the presence of ionic organic compounds led to intensive fouling and/or leakage of organics into the concentrate stream. It was concluded that to overcome these limitations it was advisable to apply electrodialysis reversal and mono-selective ion-exchange membranes

The use of polyelectrolytes in ultrafiltration of dye and surfactant solutions

Oceniono wydajność i skuteczność procesu ultrafiltracji wodnych roztworów barwników organicznych z dodatkiem substancji powierzchniowo czynnej i kationowego polielektrolitu. W badaniach wykorzystano płaskie membrany ultrafiltracyjne wykonane z regenerowanej celulozy i polietersulfonu. Membrany charakteryzowały się różnymi wartościami granicznej rozdzielczości (5 kDa, 10 kDa i 30 kDa). Proces ultrafiltracji przeprowadzono w przy ciśnieniach 0,1 MPa i 0,2 MPa w stosunku do roztworów anionowych barwników organicznych (oranżu metylowego, czerwieni indygo, czerni amidowej i czerni bezpośredniej) oraz roztworów barwników z dodatkiem surfaktantu - dodecylosiarczanu sodu (SDS) i kationowego polielektrolitu Magnofloc LT 22S. Stężenie barwników i SDS w roztworach wodnych wynosiło 100 g/m3, zaś polielektrolitu - 0,5 g/m3. Stwierdzono, że obecność kationowego polielektrolitu, jako substancji kompleksującej, przyczyniła się do polepszenia właściwości separacyjnych membran ultrafiltracyjnych (zwłaszcza polietersulfonowych) w stosunku do anionowych barwników organicznych i anionowej substancji powierzchniowo czynnej (SDS). Obecność kationowego polielektrolitu zarówno w roztworach zawierających anionowe barwniki organiczne jak i w roztworach barwników z anionową substancją powierzchniowo czynną spowodowała zmniejszenie wydajności membran ultrafiltracyjnych, zwłaszcza o umiarkowanej hydrofilowości. Przyczyną tego zjawiska było blokowanie struktury membran przez makrocząsteczkowe kompleksy typu polielektrolit-barwnik (SDS). Zjawisko to było tym bardziej widoczne, im większa była graniczna rozdzielczość membran.The effect of a cationic polyelectrolyte on the efficiency of dye and surfactant solution treatment by ultrafiltration was assessed. Flat UF membranes made of regenerated cellulose and polyethersulfone characterized by different cut-off values (5, 10, and 30 kDa) were used in the experiments. The UF process involved a pressure of 0.1 and 0.2 MPa. Permeation measurements were carried out with aqueous solutions of different anionic dyes (Methyl Orange, Indigo Carmine, Amido Black, and Direct Black) and dye solutions containing an anionic surfactant, sodium dodecyl sulphate (SDS), and/or a cationic polyelectrolyte, Magnofloc LT 22S. Dye and surfactant concentration in the experimental solutions amounted to 100 g/m3, whereas polyelectrolyte concentration was equal to 0.5 g/m3. It was found that when a cationic polyelectrolyte was present in the dye solutions or in the aqueous mixtures of an anionic dye and an anionic surfactant, this was concomitant with a decrease in membrane permeability and an increase in dye and surfactant retention (especially for polyethersulfone membranes). The observed effects can be attributed to the formation of intermolecular complexes of polycations with dye and surfactant particles. These polyelectrolyte complexes can cause membrane fouling, which is more pronounced for membranes of higher cut-off values and moderate hydrophilicity

Recovery of dyes and mineral salts from process waters by batch electrodialysis with monoanion selective ion-exchange membranes

Określono skuteczności odsalania roztworów anionowych barwników organicznych metodą elektrodializy porcjowej z wykorzystaniem membran monoanionoselektywnych (zamiast standardowych membran anionowymiennych). W badaniach zastosowano zasolone (NaCl) roztwory barwników organicznych różniących się masą cząsteczkową (327÷1060 Da). Gęstość prądu podczas elektrodializy zmieniano w zakresie 0,78÷3,91 mA/cm2. Wykazano, że skuteczność rozdziału anionowych barwników organicznych od soli mineralnych zależała od zastosowanej wartości gęstości prądu, natomiast wpływ masy cząsteczkowej barwników był niewielki. Mając na uwadze odzyskanie jak największej ilości barwników i zakładając 95% stopień odsolenia roztworu, proces elektrodializy należy prowadzić przy gęstości prądu w zakresie 1÷1,4ilim. Przy mniejszych wartościach gęstości prądu dochodziło do zjawiska sorpcji membranowej barwników, zaś przy wartościach większych barwniki przenikały przez membrany do komór koncentratu. Prowadzenie procesu elektrodializy mieszanin barwników z solami mineralnymi przy dużych wartościach gęstości prądu (>2ilim) zdecydowanie prowadziło do skrócenia czasu trwania procesu, jednakże przyczyniło się do znacznego pogorszenia jakości diluatu i zwiększenia zużycia energii.Laboratory tests were conducted to determine the efficiency of dye solution desalination by batch electrodialysis involving monoanion selective ion-exchange membranes (instead of conventional anion-exchange membranes). In the study use was made of saline (NaCl) organic dye solutions differing in molecular weight (from 327 to 1080 Da). In the course of electrodialysis current density varied from 0.78 to 3.91 mA/cm2. It was found that the efficiency of anionic organic dye separation from mineral salts was strongly influenced by the value of the current density applied, and that the influence of the dye's molecular weight was negligible. Taking into account the need of recovering the highest possible quantity of dyes, and assuming a 95% extent of desalination, it was necessary to conduct the electrodialysis process at the current density of 1-1.4ilim. When the process involved lower current density values, the dyes were adsorbed by the membranes. When the current density values were higher, the dyes penetrated through the membranes into the concentrate. At high current density (>2ilim), the duration of the electrodialysis process was noticeably shorter, but the quality of the diluate deteriorated significantly and energy demand increased